银杏叶黄酮积累相关基因克隆及查尔酮合成酶基因启动子功能研究
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摘要
为了深入研究银杏叶黄酮合成的分子机理,为今后利用生物技术手段提高银杏黄酮含量奠定基础,本文从银杏中克隆并研究了与黄酮积累相关的几个酶基因：查尔酮异构酶(GbCHI),类黄酮3'-羟化酶(F3'H),烯醇式丙酮基莽草酸3磷酸合成酶(EPSP),肉桂酰辅酶A还原酶(CCR)及查尔酮合成酶基因启动子(CHSP)序列。主要研究内容及结果如下：
(1)银杏查尔酮异构酶基因(GbCHI)的克隆、性质及表达模式的研究。利用简并PCR和RACE技术从银杏叶片中克隆得到GbCHI的cDNA序列和基因组全长。通过信息学分析发现,基因组GbCHI含有两个内含子三个外显子,GbCHI cDNA全长为926bp,含有一个735bp的开放式阅读框(ORF),编码244个氨基酸序列,预测分子量为26.29kDa,等电点为7.76。蛋白质同源序列分析表明,GbCHI与TypeⅠ型CHI同源性较高。CHIs蛋白序列进化树分析结果显示GbCHI未聚合到两类中,且分化时间早于TypeⅠ和TypeⅡ型CHI。Southern blot分析表明,GbCHI属于多基因家族；GbCHI重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测蛋白大小一致,亲和层析及Western blot分析显示,重组GbCHI且含有6xHis标签,GbCHI能在大肠杆菌中正常表达；重组GbCHI酶活性分析表明,GbCHI具有TypeⅠ型CHI的酶催化特点,即催化6'-羟基查尔酮生成(2S)-黄烷酮；RT-PCR分析显示,GbCHI基因在银杏不同组织中都有表达,但存在较大差异,只有成熟叶和雄蕊中表达量最高。CHI与银杏叶黄酮含量的年周期变化分析显示,CHI基因的转录水平与CHI的酶活性呈线性相关,相关系数为0.421；酶活性与黄酮的年周期变化之间也呈线性相关,相关系数为0.373。激素和胁迫诱导表达分析显示,虽然诱导表达模式不尽相同,但GbCHI转录水平能被UV-B、CCC、ABA、ALA和ETH诱导上调,而被GA抑制表达；GbCHI诱导表达模式与银杏叶黄酮的调控变化相一致,暗示GbCHI在银杏黄酮代谢过程中具有关键酶作用。
(2)银杏类黄酮3'-羟化酶基因(GbF3'H)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶片中克隆得到了GbF3'H的cDNA全长序列。通过信息学分析发现GbF3'H的cDNA全长为2144 bp,含有一个1671 bp的开放式阅读框(ORF),编码556个氨基酸序列。蛋白质同源序列分析表明,GbF3'H与其他物种F3'H同源性较低,而与菊苣(Cichorium intybus)同源性最高,为56.3%。同源建模分析显示GbF3'H与P450家族蛋白的三维结构及活性位点高度相似,最终定位于微粒体膜上。F3'Hs蛋白序列进化树分析结果显示GbF3'H与其他植物分化较早。Southern blot分析表明,GbF3'H属于多基因家族。GbF3'H重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测蛋白大小基本一致,亲和层析及Western blot分析显示,重组GbF3'H含有6xHis标签,GbF3'H能在大肠杆菌中正常表达。RT-PCR分析显示,GbF3'H基因在银杏不同组织中都有表达,其中雄蕊表达水平最高,其次为成熟叶。激素和胁迫诱导表达分析显示,虽然诱导表达模式不相同,但GbF3'H转录水平能被UV-B、6-BA、SA、ABA和IAA诱导上调,而伤害处理对其表达量无明显改变。GbF3'H诱导表达模式与银杏ANS基因的表达模式相似,而且其上游调控序列也发现有相关的调节单元,意味着该GbF3'H基因可能参与了银杏花色素的合成代谢。
(3)银杏烯醇式丙酮基莽草酸3磷酸合成酶(EPSPs)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶中克隆到EPSP合酶基因(GbEPSPs)的cDNA全长序列。信息学分析发现,GbEPSP的cDNA全长1403 bp,包含最大阅读框(ORF)为1035 bp,编码一个344氨基酸多肽序列。蛋白质同源序列分析表明,银杏EPSP合酶蛋白质序列与其他物种的EPSP合酶同源性较高,在81%-84%之间。进化树分析结果表明,在参试物种中GbEPSPs作为裸子植物与被子植物同聚为一类,但分歧时间相对更早。Southern blot分析表明,EPSPs基因有多个拷贝,属于一个小的多基因家族。不同组织表达分析显示,EPSPs基因在银杏的叶和果中表达量最高,其次为茎,根中表达水平最低。草甘膦处理能显著诱导银杏EPSPs基因表达量升高,紫外能上调银杏EPSPs基因表达,ABA则诱导GbEPSPs表达量先升后降；温度对GbEPSPs具有不同的诱导作用,其中42℃高温诱导最显著4 h达最大值,后又迅速降低。暗示银杏叶片中EPSPs基因在环境压力下表达量升高有可能与芳香族氨基酸向黄酮类物质的转化有关。
(4)银杏肉桂酰辅酶A还原酶基因(GbCCR)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶片中克隆得到了GbCCR的cDNA全长序列。信息学分析发现,GbCCR的cDNA全长为1178 bp,含有一个972 bp的开放式阅读框(ORF),编码323个氨基酸序列。蛋白质同源序列分析表明,GbCCR与其他物种CCR同源性相对较高,与挪威云杉(Picea abies)同源性最高,为68.6%；同源建模分析显示GbCCR序列与其他家族蛋白的三维结构及活性位点高度相似。CCRs蛋白序列进化树分析结果显示GbCCR与其他植物分化较早。Southern blot分析表明,GbCCR属于多基因家族；GbCCR重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测融合蛋白大小基本一致,亲和层析及Western blot分析显示,重组GbCCR含有6xHis标签,GbCCR能在大肠杆菌中正常表达；RT-PCR分析显示,GbCCR基因在银杏不同组织中都有表达,但与其他黄酮类代谢基因表达差异较大,其中茎和根中表达水平最高,其次为成熟叶。GA和农杆菌诱导表达分析显示,虽然二者都能诱导基因表达,但GA能够明显诱导GbCCR转录水平上调,而农杆菌侵染处理对其表达量无明显改变。GbCCR诱导表达模式显示,该基因可能主要参与组织及细胞壁中木质素的合成,而与抗病虫害无明显关系。
(5)银杏查尔酮合成酶基因启动子(GbCHSP)的调控元件及功能分析。通过染色体步移方法从银杏基因组中克隆到查尔酮合成酶基因(GbCHS)翻译起始位点上游1711bp的启动子序列。生物信息学分析表明,该启动子片段中存在多个顺式作用元件,包括紫外／蓝光响应单元、植物激素响应单元、真菌诱导元件、MYB结合位点、TATA-box和CAAT-box等。亚克隆了GbCHS转录起始位点上游1402 bp序列,将其与GUS基因构建融合表达载体pBI121+CHSP,以pBI121-35s作为负对照,通过农杆菌(LBA4404)介导法分别转入烟草。结果表明,银杏CHS启动子序列能驱动GUS基因在烟草中的表达,表达具有组织差异性。其中叶片和茎中表达量较高；在诱导芽及愈伤组织中亦具有较高表达水平。结果说明,CHSP具有基本启动子功能,但表现出空间上的表达差异性
In order to study biosynthetic mechanism of flavonoids in G.biloba and develop biotechnology of increasing the content of the flavonoids, several key genes such as GbCHI, GbF3'H, GbCCR, GbEPSPs and GbCHSp which are involved in the flavonoids accumulation process have been cloned and studied in this paper. The main results are as follows:
(1) Molecular cloning, characterization and expression models of chalcone isomerase gene from Ginkgo biloba. A full-length cDNA and genomic DNA of chalcone isomerase gene were isolated from Ginkgo biloba L. by using PCR and RACE technologies. Compared with the cDNA sequence, the genomic sequence contain two intron and three extron. The coding region of the gene is 735 bp long, and its deduced protein consists of 244 amino acids with a predicted molecular mass of 26.29 kDa and a pI of 7.76. Protein sequence analysis reveals that GbCHI had closer relationship with TypeⅠCHIs. Phylogenetic tree analysis revealed that GbCHI does not belong to TypeⅠor typeⅡgroup. Southern blot analysis indicated that GbF3'H belonged to a multi-gene family. The results of prokaryotic expression showed that the product of recombinant GbCHI protein was in accordance with the anticipation. The vitro enzyme activity assay by HPLC indicated that recombinant GbCHI protein could catalyze the formation the (2S) naringenin from 6'-hydroxychalcone. The expression analysis by RT-PCR showed that GbCHI constitutively expressed in all the tested tissues, and had tissue specific manner in G.biloba. GbCHI was also found to be up-regulated by UV, ALA, ETH, ABA, CCC, and be down-regulated by GA. Correlation analysis between CHI activity and flavonoid accumulation during gingkgo leaf growth indicated that GbCHI might be the rate-limiting enzyme in the biosynthesis pathway of flavonoids in ginkgo leaves. Results of quantitative RT-PCR analysis showed that CHI activity correlated with the transcription level of change in CHI gene, suggesting CHI gene as the specific key gene regulating flavonoid accumulation in ginkgo.
(2) Molecular cloning, characterization and expression models of Flavonoid 3'-hydroxylase gene from G.biloba. Flavonoid 3'-hydroxylase (GbF3'H), is a member of the P450 superfamily, which catalyzes monooxygenase reactions dependent upon NADPH and O2. In the flavonoid pathway F3'H hydroxylates the 3'-position of the B ring of naringenin and dihydrokaempferol to generate eriodictyol and dihydroquercetin, respectively which are important intermediates for biosynthesis of anthocyanins and proanthocyanidins, major coloration substances of flowers and seed coat. The full-length cDNA sequences of F3'H gene (designated as GbF3'H) were isolated from G.biloba for the first time. The full-length cDNA of GbF3'H contains a 1671 bp open reading frame (ORF) encoding a 556 amino acid protein. The 5'flanking region of GbF3'H was isolated by genome walking method, and some main cis-acting elements including TATA box and stress-responsiveness elements were predicted and analyzed. The deduced GbF3'H protein showed low identities to other plant F3'Hs, but had closer relationship with Cichorium intybus and shares 56.3% homology.3D structure modeling showed that GbF3'H bears all conserved motifs featured and have high similarity with P450s. Phylogenetic tree analysis revealed that the divergence time of GbF3'H from other F3'Hs is earlier. Southern blot analysis indicated that GbF3'H belonged to a multi-gene family. The expression analysis by RT-PCR showed that GbF3'H expressed in a tissue-specific manner in G.biloba, with the highest level in stamen and next in mature leaves. GbF3'H was also found to be up-regulated by the five tested abiotic stresses:UV-B、6-BA、SA、ABA and IAA, but non significant effect to wounding. Function analysis suggest that GbF3'H is a functional enzyme within the anthocyanidin and flavonol biosynthetic pathway.
(3) Molecular cloning, characterization and expression models of EPSP synthase Gene From Ginkgo biloba L. The full-length cDNA sequences of EPSP synthase gene (designated as GbEPSPs) were isolated from G.biloba by using PCR and RACE technologies for the first time. The full-length cDNA of GbEPSPs is 1404bp and contains a 1035 bp open reading frame (ORF) encoding a 344 amino acid peptide sequence. Protein sequence analysis reveals that GbEPSPs had close relationship with other EPSPs and shares 81%-84% homology. Phylogenetic tree analysis showed that GbEPSPs as a gymnosperm and other angiosperm EPSPs are clustered to one monophyletic group, but the divergence time is early. RT-PCR analysis showed that GbEPSPs expressed in leaves, stems, roots and fruits, and had the highest expression in leaves and fruits, the next in stems, the least in roots. The expression of GbEPSPs could be induced by glyphosate and UV-B. ABA could improve the expression of GbEPSPs first, but deduce later. Different temperature treatments have different effects in the content of GbEPSP gene, and the highest expression at 42℃for 4h.
(4) Molecular cloning, characterization and expression models of Cinnamoyl-CoA Reductase gene from G.biloba. Cinnamoyl-CoA Reductase (CCR, EC 1.2.1.44) catalyses the first step of the lignin pathway. The full-length cDNA sequences of CCR gene (designated as GbCCR) were isolated from G.biloba for the first time. The full-length cDNA of GbCCR is 1178 bp long and contains a 972 bp open reading frame (ORF) encoding a 323 amino acid protein. The deduced GbCCR protein showed high identities to other plant CCRs, and had closer relationship with Picea abies shares 56.3% homology. They both contain a common signature which is thought to be involved in the catalytic site of CCR. Phylogenetic tree analysis revealed that GbCCR shared the same ancestor with other CCRs, but the divergence time is early. Southern blot analysis indicated that GbCCR belonged to a multi-gene family. The expression analysis by RT-PCR showed that GbCCR had tissue specific manner in G.biloba, and the highest expression in stems and roots, the next in mature leaves, which were great difference with other flavones biosynthetic pathway gene. GbCCR was also found to be significant up-regulated by GA, but agrobacterium treatment had no significant change. The high level of GbCCR gene expression along the stalk suggests that the corresponding enzyme is probably involve in constitutive lignification.
(5) Regulatory Element and Function Analysis of Chalcone Synthase Gene Promoter from Ginkgo biloba L. The regulative sequence (1711 bp) of chalcone synthase gene promoter (CHSP) from Ginkgo biloba L. was cloned by genomic walking. In bioinformatic analysis of sequence suggested that the sequence contained several typial cis-acting elements, including UV/blue light responsive elements, Phytohormone responsive elements, fungal elicitor responsive elements, MYB binding site, TATA-box and CAAT-box. A 1402 bp promoter sequence upstream 5'of translation start site of GbCHS were cloned and designated as GbCHSP, respectively. pBI121+CHSP and pBI121-35s were constructed and transformed into tobacco by LBA4404. These result showed that pBI121 and pBI121+CHSP both could drive the transient expression of GUS in tobacco and pBI121+CHSP expressed differentially in root, stem and leaf tissues of tobacco. Our discoveries will be help to understand the transcriptional regulatory mechanism on GbCHS expression and accumulation flavonoids.
(1)银杏查尔酮异构酶基因(GbCHI)的克隆、性质及表达模式的研究。利用简并PCR和RACE技术从银杏叶片中克隆得到GbCHI的cDNA序列和基因组全长。通过信息学分析发现,基因组GbCHI含有两个内含子三个外显子,GbCHI cDNA全长为926bp,含有一个735bp的开放式阅读框(ORF),编码244个氨基酸序列,预测分子量为26.29kDa,等电点为7.76。蛋白质同源序列分析表明,GbCHI与TypeⅠ型CHI同源性较高。CHIs蛋白序列进化树分析结果显示GbCHI未聚合到两类中,且分化时间早于TypeⅠ和TypeⅡ型CHI。Southern blot分析表明,GbCHI属于多基因家族；GbCHI重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测蛋白大小一致,亲和层析及Western blot分析显示,重组GbCHI且含有6xHis标签,GbCHI能在大肠杆菌中正常表达；重组GbCHI酶活性分析表明,GbCHI具有TypeⅠ型CHI的酶催化特点,即催化6'-羟基查尔酮生成(2S)-黄烷酮；RT-PCR分析显示,GbCHI基因在银杏不同组织中都有表达,但存在较大差异,只有成熟叶和雄蕊中表达量最高。CHI与银杏叶黄酮含量的年周期变化分析显示,CHI基因的转录水平与CHI的酶活性呈线性相关,相关系数为0.421；酶活性与黄酮的年周期变化之间也呈线性相关,相关系数为0.373。激素和胁迫诱导表达分析显示,虽然诱导表达模式不尽相同,但GbCHI转录水平能被UV-B、CCC、ABA、ALA和ETH诱导上调,而被GA抑制表达；GbCHI诱导表达模式与银杏叶黄酮的调控变化相一致,暗示GbCHI在银杏黄酮代谢过程中具有关键酶作用。
(2)银杏类黄酮3'-羟化酶基因(GbF3'H)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶片中克隆得到了GbF3'H的cDNA全长序列。通过信息学分析发现GbF3'H的cDNA全长为2144 bp,含有一个1671 bp的开放式阅读框(ORF),编码556个氨基酸序列。蛋白质同源序列分析表明,GbF3'H与其他物种F3'H同源性较低,而与菊苣(Cichorium intybus)同源性最高,为56.3%。同源建模分析显示GbF3'H与P450家族蛋白的三维结构及活性位点高度相似,最终定位于微粒体膜上。F3'Hs蛋白序列进化树分析结果显示GbF3'H与其他植物分化较早。Southern blot分析表明,GbF3'H属于多基因家族。GbF3'H重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测蛋白大小基本一致,亲和层析及Western blot分析显示,重组GbF3'H含有6xHis标签,GbF3'H能在大肠杆菌中正常表达。RT-PCR分析显示,GbF3'H基因在银杏不同组织中都有表达,其中雄蕊表达水平最高,其次为成熟叶。激素和胁迫诱导表达分析显示,虽然诱导表达模式不相同,但GbF3'H转录水平能被UV-B、6-BA、SA、ABA和IAA诱导上调,而伤害处理对其表达量无明显改变。GbF3'H诱导表达模式与银杏ANS基因的表达模式相似,而且其上游调控序列也发现有相关的调节单元,意味着该GbF3'H基因可能参与了银杏花色素的合成代谢。
(3)银杏烯醇式丙酮基莽草酸3磷酸合成酶(EPSPs)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶中克隆到EPSP合酶基因(GbEPSPs)的cDNA全长序列。信息学分析发现,GbEPSP的cDNA全长1403 bp,包含最大阅读框(ORF)为1035 bp,编码一个344氨基酸多肽序列。蛋白质同源序列分析表明,银杏EPSP合酶蛋白质序列与其他物种的EPSP合酶同源性较高,在81%-84%之间。进化树分析结果表明,在参试物种中GbEPSPs作为裸子植物与被子植物同聚为一类,但分歧时间相对更早。Southern blot分析表明,EPSPs基因有多个拷贝,属于一个小的多基因家族。不同组织表达分析显示,EPSPs基因在银杏的叶和果中表达量最高,其次为茎,根中表达水平最低。草甘膦处理能显著诱导银杏EPSPs基因表达量升高,紫外能上调银杏EPSPs基因表达,ABA则诱导GbEPSPs表达量先升后降；温度对GbEPSPs具有不同的诱导作用,其中42℃高温诱导最显著4 h达最大值,后又迅速降低。暗示银杏叶片中EPSPs基因在环境压力下表达量升高有可能与芳香族氨基酸向黄酮类物质的转化有关。
(4)银杏肉桂酰辅酶A还原酶基因(GbCCR)的克隆、性质及表达模式研究。利用简并PCR和RACE技术从银杏叶片中克隆得到了GbCCR的cDNA全长序列。信息学分析发现,GbCCR的cDNA全长为1178 bp,含有一个972 bp的开放式阅读框(ORF),编码323个氨基酸序列。蛋白质同源序列分析表明,GbCCR与其他物种CCR同源性相对较高,与挪威云杉(Picea abies)同源性最高,为68.6%；同源建模分析显示GbCCR序列与其他家族蛋白的三维结构及活性位点高度相似。CCRs蛋白序列进化树分析结果显示GbCCR与其他植物分化较早。Southern blot分析表明,GbCCR属于多基因家族；GbCCR重组蛋白大肠杆菌表达显示,其蛋白大小与cDNA序列预测融合蛋白大小基本一致,亲和层析及Western blot分析显示,重组GbCCR含有6xHis标签,GbCCR能在大肠杆菌中正常表达；RT-PCR分析显示,GbCCR基因在银杏不同组织中都有表达,但与其他黄酮类代谢基因表达差异较大,其中茎和根中表达水平最高,其次为成熟叶。GA和农杆菌诱导表达分析显示,虽然二者都能诱导基因表达,但GA能够明显诱导GbCCR转录水平上调,而农杆菌侵染处理对其表达量无明显改变。GbCCR诱导表达模式显示,该基因可能主要参与组织及细胞壁中木质素的合成,而与抗病虫害无明显关系。
(5)银杏查尔酮合成酶基因启动子(GbCHSP)的调控元件及功能分析。通过染色体步移方法从银杏基因组中克隆到查尔酮合成酶基因(GbCHS)翻译起始位点上游1711bp的启动子序列。生物信息学分析表明,该启动子片段中存在多个顺式作用元件,包括紫外／蓝光响应单元、植物激素响应单元、真菌诱导元件、MYB结合位点、TATA-box和CAAT-box等。亚克隆了GbCHS转录起始位点上游1402 bp序列,将其与GUS基因构建融合表达载体pBI121+CHSP,以pBI121-35s作为负对照,通过农杆菌(LBA4404)介导法分别转入烟草。结果表明,银杏CHS启动子序列能驱动GUS基因在烟草中的表达,表达具有组织差异性。其中叶片和茎中表达量较高；在诱导芽及愈伤组织中亦具有较高表达水平。结果说明,CHSP具有基本启动子功能,但表现出空间上的表达差异性
In order to study biosynthetic mechanism of flavonoids in G.biloba and develop biotechnology of increasing the content of the flavonoids, several key genes such as GbCHI, GbF3'H, GbCCR, GbEPSPs and GbCHSp which are involved in the flavonoids accumulation process have been cloned and studied in this paper. The main results are as follows:
(1) Molecular cloning, characterization and expression models of chalcone isomerase gene from Ginkgo biloba. A full-length cDNA and genomic DNA of chalcone isomerase gene were isolated from Ginkgo biloba L. by using PCR and RACE technologies. Compared with the cDNA sequence, the genomic sequence contain two intron and three extron. The coding region of the gene is 735 bp long, and its deduced protein consists of 244 amino acids with a predicted molecular mass of 26.29 kDa and a pI of 7.76. Protein sequence analysis reveals that GbCHI had closer relationship with TypeⅠCHIs. Phylogenetic tree analysis revealed that GbCHI does not belong to TypeⅠor typeⅡgroup. Southern blot analysis indicated that GbF3'H belonged to a multi-gene family. The results of prokaryotic expression showed that the product of recombinant GbCHI protein was in accordance with the anticipation. The vitro enzyme activity assay by HPLC indicated that recombinant GbCHI protein could catalyze the formation the (2S) naringenin from 6'-hydroxychalcone. The expression analysis by RT-PCR showed that GbCHI constitutively expressed in all the tested tissues, and had tissue specific manner in G.biloba. GbCHI was also found to be up-regulated by UV, ALA, ETH, ABA, CCC, and be down-regulated by GA. Correlation analysis between CHI activity and flavonoid accumulation during gingkgo leaf growth indicated that GbCHI might be the rate-limiting enzyme in the biosynthesis pathway of flavonoids in ginkgo leaves. Results of quantitative RT-PCR analysis showed that CHI activity correlated with the transcription level of change in CHI gene, suggesting CHI gene as the specific key gene regulating flavonoid accumulation in ginkgo.
(2) Molecular cloning, characterization and expression models of Flavonoid 3'-hydroxylase gene from G.biloba. Flavonoid 3'-hydroxylase (GbF3'H), is a member of the P450 superfamily, which catalyzes monooxygenase reactions dependent upon NADPH and O2. In the flavonoid pathway F3'H hydroxylates the 3'-position of the B ring of naringenin and dihydrokaempferol to generate eriodictyol and dihydroquercetin, respectively which are important intermediates for biosynthesis of anthocyanins and proanthocyanidins, major coloration substances of flowers and seed coat. The full-length cDNA sequences of F3'H gene (designated as GbF3'H) were isolated from G.biloba for the first time. The full-length cDNA of GbF3'H contains a 1671 bp open reading frame (ORF) encoding a 556 amino acid protein. The 5'flanking region of GbF3'H was isolated by genome walking method, and some main cis-acting elements including TATA box and stress-responsiveness elements were predicted and analyzed. The deduced GbF3'H protein showed low identities to other plant F3'Hs, but had closer relationship with Cichorium intybus and shares 56.3% homology.3D structure modeling showed that GbF3'H bears all conserved motifs featured and have high similarity with P450s. Phylogenetic tree analysis revealed that the divergence time of GbF3'H from other F3'Hs is earlier. Southern blot analysis indicated that GbF3'H belonged to a multi-gene family. The expression analysis by RT-PCR showed that GbF3'H expressed in a tissue-specific manner in G.biloba, with the highest level in stamen and next in mature leaves. GbF3'H was also found to be up-regulated by the five tested abiotic stresses:UV-B、6-BA、SA、ABA and IAA, but non significant effect to wounding. Function analysis suggest that GbF3'H is a functional enzyme within the anthocyanidin and flavonol biosynthetic pathway.
(3) Molecular cloning, characterization and expression models of EPSP synthase Gene From Ginkgo biloba L. The full-length cDNA sequences of EPSP synthase gene (designated as GbEPSPs) were isolated from G.biloba by using PCR and RACE technologies for the first time. The full-length cDNA of GbEPSPs is 1404bp and contains a 1035 bp open reading frame (ORF) encoding a 344 amino acid peptide sequence. Protein sequence analysis reveals that GbEPSPs had close relationship with other EPSPs and shares 81%-84% homology. Phylogenetic tree analysis showed that GbEPSPs as a gymnosperm and other angiosperm EPSPs are clustered to one monophyletic group, but the divergence time is early. RT-PCR analysis showed that GbEPSPs expressed in leaves, stems, roots and fruits, and had the highest expression in leaves and fruits, the next in stems, the least in roots. The expression of GbEPSPs could be induced by glyphosate and UV-B. ABA could improve the expression of GbEPSPs first, but deduce later. Different temperature treatments have different effects in the content of GbEPSP gene, and the highest expression at 42℃for 4h.
(4) Molecular cloning, characterization and expression models of Cinnamoyl-CoA Reductase gene from G.biloba. Cinnamoyl-CoA Reductase (CCR, EC 1.2.1.44) catalyses the first step of the lignin pathway. The full-length cDNA sequences of CCR gene (designated as GbCCR) were isolated from G.biloba for the first time. The full-length cDNA of GbCCR is 1178 bp long and contains a 972 bp open reading frame (ORF) encoding a 323 amino acid protein. The deduced GbCCR protein showed high identities to other plant CCRs, and had closer relationship with Picea abies shares 56.3% homology. They both contain a common signature which is thought to be involved in the catalytic site of CCR. Phylogenetic tree analysis revealed that GbCCR shared the same ancestor with other CCRs, but the divergence time is early. Southern blot analysis indicated that GbCCR belonged to a multi-gene family. The expression analysis by RT-PCR showed that GbCCR had tissue specific manner in G.biloba, and the highest expression in stems and roots, the next in mature leaves, which were great difference with other flavones biosynthetic pathway gene. GbCCR was also found to be significant up-regulated by GA, but agrobacterium treatment had no significant change. The high level of GbCCR gene expression along the stalk suggests that the corresponding enzyme is probably involve in constitutive lignification.
(5) Regulatory Element and Function Analysis of Chalcone Synthase Gene Promoter from Ginkgo biloba L. The regulative sequence (1711 bp) of chalcone synthase gene promoter (CHSP) from Ginkgo biloba L. was cloned by genomic walking. In bioinformatic analysis of sequence suggested that the sequence contained several typial cis-acting elements, including UV/blue light responsive elements, Phytohormone responsive elements, fungal elicitor responsive elements, MYB binding site, TATA-box and CAAT-box. A 1402 bp promoter sequence upstream 5'of translation start site of GbCHS were cloned and designated as GbCHSP, respectively. pBI121+CHSP and pBI121-35s were constructed and transformed into tobacco by LBA4404. These result showed that pBI121 and pBI121+CHSP both could drive the transient expression of GUS in tobacco and pBI121+CHSP expressed differentially in root, stem and leaf tissues of tobacco. Our discoveries will be help to understand the transcriptional regulatory mechanism on GbCHS expression and accumulation flavonoids.
引文
[1]王燕,程水源,汪琼.银杏栽培学[M].武汉：湖北人民出版社,2007：
[2]程水源,顾曼如,束怀瑞.银杏叶黄酮研究进展[J].林业科学.2000,36(6)：110-115.
[3]毕殉,卢嘉文,蔡东联.银杏及叶中黄酮类化合物生理功效的研究进展[J].武警医学.2004,15(6)：458-459.
[4]李勇,刘新民.银杏及叶中黄酮类化合物生理功效的研究进展[J].食品科技.2001,(5)：72-73.
[5]Cook N, Samman S. Flavonoids-chemistry, metabolism, cardioprotective effects, and dietary sources[J]. The Journal of nutritional biochemistry.1996,7(2):66-76.
[6]Winkel-Shirley B. Biosynthesis of flavonoids and effects of stress [J]. Current opinion in plant biology.2002,5(3):218-223.
[7]Tahara S, Ibrahim R K. Prenylated isoflavonoids-an update[J]. Phytochemistry.1995,38(5): 1073-1094.
[8]Dooner H, Robbins T, Jorgensen R. Genetic and developmental control of anthocyanin biosynthesis[J]. Annual Review of Genetics.1991,25(1):173-199.
[9]Koes R, Quattrocchio F, Mol J. The flavonoid biosynthetic pathway in plants:function and evolution[J]. BioEssays.1994,16(2):123-132.
[10]Dixon R, Paiva N. Stress-induced phenylpropanoid metabolism[J]. The Plant Cell.1995,7(7): 1085.
[11]Schijlen E, Ric de Vos C, van Tunen A, et al. Modification of flavonoid biosynthesis in crop plants[J]. Phytochemistry.2004,65(19):2631-2648.
[12]梁立兴.中国当代银杏大全[M].1993：1-41.
[13]程水源.银杏叶化学成分与药用研究进展[J].中草药.1998,(29)：16-18.
[14]邵继平,王伯初.银杏叶提取物药用价值的研究进展[J].重庆大学学报：自然科学版.2003,26(1)：130-134.
[15]古勇,李安明.类黄酮生物活性的研究进展[J].应用与环境生物学报.2006,12(2)：283-286.
[16]Hasler A, Gross G A, Meier B, et al. Complex flavonol glycosides from the leaves of Ginkgo biloba[J]. Phytochemistry.1992,31(4):1391.
[17]夏晓晖,张宇,郗砚彬等.银杏叶化学成分研究进展[J].中国实验方剂学杂志.2009,15(9)：100-104.
[18]唐于平,王颖.银杏叶中的黄酮醇苷类成分[J].药学学报.2000,35(5)：363-366.
[19]唐于平,楼凤昌.银杏叶中黄酮类成分的研究[J].中国药学杂志.2001,36(4)：231-233.
[20]Dixon R A, Steele C L. Flavonoids and isoflavonoids-a gold mine for metabolic engineering[J]. Trends in plant science.1999,4(10):394-400.
[21]Holton T A, Cornish E C. Genetics and biochemistry of anthocyanin biosynthesis[J]. The Plant Cell. 1995,7(7):1071.
[22]Tsai C J, El Kayal W, Harding S A. Populus, the new model system for investigating phenylpropanoid complexity [J]. Int J Appl Sci Eng.2006,4:221-233.
[23]Pla J, Ville A, Pacheco H. Biogenesis of plant pigments.1. Comparative study of the incorporation of 1,2-14C shikimic and 3-14C trans-cinnamic acids in two anthocyanic pigment derivatives of delphinidine and cyanidine[J]. Bull Soc Chim Biol.1967,49(4):395-413.
[24]Zaprometov M N, Bukhlaeva V I. Efficiency of use of various C14-precursors for the biosynthesis of flavonoids in the tea plant[J]. Biokhimiia.36(2):270.
[25]Zenk M H. Biosynthese von vanillin in Vanilla planifolia Andr[J]. Z. Pflanzenphysiol.1965,53: 404-414.
[26]Stafford H A. Possible multi-enzyme complexes regulating the formation of C6-C3 phenolic compounds and lignins in higher plants[J]. Rec. Adv. Phytochem.1974,8:53-79.
[27]Stafford H. Flavonoid evolution:an enzymic approach[J]. Plant Physiology.1991,96(3):680.
[28]方从兵,宛晓春,江昌俊.黄酮类化合物生物合成的研究进展(综述)[J].安徽农业大学学报,2005,32(4)：498-504.
[29]Christendat D, Saridakis V C, Turnbull J L. Use of Site-Directed Mutagenesis To Identify Residues Specific for Each Reaction Catalyzed by Chorismate Mutase-Prephenate Dehydrogenase from Escherichia coli[J]. Biochemistry.1998,37(45):15703-15712.
[30]Koch G L E, Shaw D C, Gibson F. Tyrosine biosynthesis in Aerobacter aerogenes:purification and properties of chorismate mutase-prephenate dehydrogenase[J]. Biochimica et Biophysica Acta (BBA)-Enzymology.1970,212(3):375-386.
[31]Stenmark S L, Pierson D L, Jensen R A, et al. Blue-green bacteria synthesise L-tyrosine by the pretyrosine pathway[J]. Nature (Lond).1974,247:290-292.
[32]Kim K H, Petersen M. Cloning and functional expression of hydroxyphenylpyruvate dioxygenase from Coleus blumei[J]. Phytochemistry.1997,45:1165-1172.
[33]petersen M, Simmonds M S J. Molecules of interest:rosmarinic acid[J]. Phytochemistry.2003, 62(2):121-125.
[34]孙视,刘晚苟.生态条件对银杏叶黄酮积累的影响[J].植物资源与环境.1998,7(3)：1-7.
[35]王义强.银杏枝叶生长规律及两次采叶试验研究[J].经济林研究.1995,13(4)：20-22.
[36]Lo S, Nicholson R. Reduction of light-induced anthocyanin accumulation in inoculated sorghum mesocotyls. Implications for a compensatory role in the defense response[J]. Plant Physiology. 1998,116(3):979.
[37]陈学森,张艳敏.叶用银杏资源评价及选优的研究[J].园艺学报.1997,24(3)：215-219.
[38]王英强,梁红.广东产银杏叶总黄酮含量变化[J].中药材.2001,24(4)：247-248.
[39]邢世岩,吴德军.叶用银杏种源,性别及无性系的因子和聚类分析[J].中南林学院学报.2000,20(2)：26-31.
[40]汪贵斌,曹福亮,方升佐等.银杏叶用园建园材料选择的研究[J].林业科学.2000,36(4)：26-31.
[41]刘德军,佘远国,汪鹏等.叶用银杏品种产量性状选择研究[J].经济林研究.2002,20(1)：30-31.
[42]王凌晖,曹福亮,汪贵斌等.银杏优质丰产园建园技术的研究进展[J].西北林学院学报.2005.20(1)：102-106.
[43]陈学森,章文才,邓秀新.树龄及季节对银杏叶黄酮与萜内酯含量的影响fJ].果树科学.1997,14(4)：226-229.
[44]程水源.影响银杏叶黄酮形成的主要因子及调控技术的研究[D].泰安：山东农业大学.2001
[45]Brinkman J, Boerner R. Nitrogen fertilization effects on foliar nutrient dynamics and autumnal resorption in maidenhair tree(Gingko biloba L.)[J]. Journal of Plant Nutrition.1994,17(2): 433-443.
[46]魏刚,赵洪亮.银杏不同营养器官中营养元素含量季节动态的研究[J].北京林业大学学报.1999,21(1)：96-99.
[47]王燕,程水源.提高银杏叶黄酮含量的调控措施[J].湖北农业科学.2002.40(5)：103-105.
[48]程水源,王燕,费永俊等.提高银杏叶黄酮含量的措施及其调控机理的研究[J].果树学报.2004,21(2)：116-119.
[49]冷平生,苏淑钗.光强与光质对银杏光合作用及黄酮苷与萜类内酯含量的影响[J].植物资源与环境学报.2002,11(1)：1-4.
[50]何丙辉,钟章成.不同光强与干旱胁迫对银杏枝叶构件生长的影响[J].广西师范大学学报：自然科学版.2005,23(3)：66-69.
[51]谢宝东,王华田.土壤水分含量对银杏叶黄酮和内酯含量的影响[J].山东林业科技.2002,(4)：1-3.
[52]景茂,曹宝亮,汪贵斌等.土壤水分含量对银杏生长及生物量分配的影响[J].南京林业大学学报：自然科学版.2005,29(3)：5-8.
[53]刘卫红,程水源.光照及机械损伤对银杏叶苯丙氨酸解氨酶活性的影响[J].湖北农业科学.2003,(3)：73-75.
[54]王燕,刘卫红,杜何为等.底物,末端产物对离体银杏叶苯丙氨酸解氨酶活性的影响[J].果树学报.2004,21(5)：443-446.
[55]王燕,李琳玲,许锋等.金属离子对盆栽银杏叶PAL酶活及黄酮含量的影响[J].南京林业大学学报：自然科学版.2007,31(2)：68-72.
[56]Xu F, Cheng S, Cheng S, et al. Time course of expression of chalcone synthase gene in Ginkgo biloba[J]. Plant Physiol. Mol. Biol.2007,33(4):309-317.
[57]Xu F, Cai R, Cheng S, et al. Molecular cloning, characterization and expression of phenylalanine ammonia-lyase gene from Ginkgo biloba[J]. African Journal of Biotechnology.2008,7(6): 721-729.
[58]Pang Y, Shen G, Wu W, et al. Characterization and expression of chalcone synthase gene from Ginkgo biloba[J]. Plant Science.2005,168(6):1525-1531.
[59]程水源,陈昆松,刘卫红等.植物苯丙氨酸解氨酶基因的表达调控与研究展望[J].果树学报.2003,20(5)：351-357.
[60]刘卫红.银杏叶苯丙氨酸解氨酶特性及其对叶黄酮含量调控的研究[D].武汉：华中农业大学, 2003
[61]Muir S R, Collins, G. J.,Robinson, S.,Hughes, S.,Bovy, A., Ric De Vos C H, van Tunen A J, et al. Overexpression of petunia chalcone isomerase in tomato results in fruit containing increased levels of flavonols[J]. Nat Biotechnol.2001,19(5):470-474.
[62]Li F X, Jin Z P, Zhao D X, et al. Overexpression of the Saussurea medusa chalcone isomerase gene in S. involucrata hairy root cultures enhances their biosynthesis of apigenin[J]. Phytochemistry. 2006,67(6):553-560.
[63]Harborne J, Williams C. Anthocyanins and other flavonoids[J]. Natural product reports.2001, 18(3):310-333.
[64]Ju Z, Liu C, Yuan Y. Activities of chalcone synthase and UDPGal:flavonoid-3-o-glycosy-ltransferase in relation to anthocyanin synthesis in apple[J]. Scientia Horticulturae.1995,63(3-4): 175-185.
[65]Welford R, Clifton I, Turnbull J, et al. Structural and mechanistic studies on anthocyanidin synthase catalysed oxidation of flavanone substrates:the effect of C-2 stereochemistry on product selectivity and mechanism[J]. Organic & Biomolecular Chemistry.2005,3(17):3117-3126.
[66]许锋.银杏GbPAL和GbANS基因的克隆与表达及ALA对类黄酮含量的影响[D].泰安：山东农业大学,2008
[67]Cosio E, McClure J. Kaempferol glycosides and enzymes of flavonol biosynthesis in leaves of a soybean strain with low photosynthetic rates[J]. Plant Physiology.1984,74(4):877.
[68]Saure M C. External control of anthocyanin formation in apple[J]. Scientia Horticulturae.1990, 42(3):181-218.
[69]El-Kereamy A, Chervin C, Roustan J, et al. Exogenous ethylene stimulates the long-term expression of genes related to anthocyanin biosynthesis in grape berries[J]. Physiologia Plantarum. 2003,119(2):175-182.
[70]Blankenship S, Unrath R. PAL and ethylene content during maturation of red and golden delicious apples[J]. Phytochemistry.1988,27(4):1001-1002.
[71]Awad M, de Jager A. Formation of flavonoids, especially anthocyanin and chlorogenic acid in [] Jonagold'apple skin:influences of growth regulators and fruit maturity [J]. Scientia Horticulturae. 2002,93(3-4):257-266.
[72]Hiratsuka S, Onodera H, Kawai Y, et al. ABA and sugar effects on anthocyanin formation in grape berry cultured in vitro [J]. Scientia Horticulturae.2001,90(1-2):121-130.
[73]刘叔倩,郑俊华.展望分子生物学技术银否研究中的应用[J].国外医药：植物约分册.1999,14(1)：4-6.
[74]Tulecke W. A Haploid Tissue Culture from the Female Gametophyte of Ginkgo biloba L[J]. Nature. 1964,203:94-95.
[75]Laurain D, Chenieux J, Tremouillaux-Guiller J. Direct embryogenesis from female haploid protoplasts of Ginkgo biloba L., a medicinal woody species[J]. Plant Cell Reports.1993,12(11): 656-660.
[76]张广辉,陈春秋.银杏离体培养生产次生代谢物研究进展[J].北京林业大学学报.2002,24(4)：130-134.
[77]陈学森,邓秀新,章文才.银杏组织培养与黄酮生产的研究[J].中国农业科学.1997,30(6)：55-60.
[78]陈学森,邓秀新.培养基及培养条件对银杏愈伤组织黄酮产量的影响[J].园艺学报.1997,24(4)：373-377.
[79]房建军,阙国宁.银杏愈伤组织生长和黄酮类化合物积累的关系[J].林业科学研究.1998,11(2)：124-129.
[80]孙天恩,李根保.银杏发根克隆及其培养技术研究进展[J].中草药.1998,29(增)：25-28.
[81]Winkel-Shirley B. Flavonoid biosynthesis. A colorful model for genetics, biochemistry, cell biology, and biotechnology[J]. Plant Physiology.2001,126(2):485-493.
[82]陈新,万德光,严铸云等.银杏苯丙氨酸解氨酶(PAL)的基因克隆[J].成都中医药大学学报.2004,27(1)：32-34.
[83]Pang Y, Shen G A, Liu C, et al. Molecular Cloning and Sequence Analysis of a Novel Chalcone Synthase cDNA from Ginkgo biloba[J]. Mitochondrial DNA.2004,15(4):283-290.
[84]Shen G, Pang Y, Wu W, et al. Cloning and characterization of a flavanone 3-hydroxylase gene from Ginkgo biloba[J]. Bioscience reports.2006,26(1):19-29.
[85]Shen G, Pang Y, Wu W, et al. Isolation and characterization of a putative anthocyanidin reductase gene from Ginkgo biloba[J]. Journal of plant physiology.2006,163(2):224-227.
[86]庞永珍.银杏黄酮和萜类化合物生物合成途径中重要相关基因的克隆和研究[D].上海：复旦大学,2005
[87]Bovy A, De Vos R, Kemper M, et al. High-flavonol tomatoes resulting from the heterologous expression of the maize transcription factor genes LC and C1[J]. The Plant Cell Online.2002, 14(10):2509.
[88]Verhoeyen M E, Bovy A, Collins G, et al. Increasing antioxidant levels in tomatoes through modification of the flavonoid biosynthetic pathway[J]. J. Exp. Bot.2002,53(377):2099-2106.
[89]Colliver S, Bovy A, Collins G, et al. Improving the nutritional content of tomatoes through reprogramming their flavonoid biosynthetic pathway[J]. Phytochemistry Reviews.2002,1(1): 113-123.
[90]Aharoni A, De Vos C, Wein M, et al. The strawberry FaMYB1 transcription factor suppresses anthocyanin and flavonol accumulation in transgenic tobacco[J]. The Plant Journal.2001,28(3): 319-332.
[91]路静,赵华燕,何奕昆等.高等植物启动子及其应用研究进展[J].自然科学进展.2004,14(8)：856-862.
[92]朱玉贤,李毅.现代分子生物学[M].北京：高等教育出版社,1997：17—59.
[93]夏江东,程在全,吴渝生等.高等植物启动子功能和结构研究进展[J].云南农业大学学报.2006,21(1)：7-14.
[94]张春晓,王文棋,蒋湘宁等.植物基因启动子研究进展[J].遗传学报.2004,31(12)：1455-1464.
[95]Zhang C X, Wang W Q, Jiang X N, et al. Review on plant gene promoters[J]. Acta genetica Sinica. 2004,31(12):1455-1464.
[96]Carey M F, Smale S T. Transcriptional regulation in eukaryotes:concepts, strategies, and techniques[M]. Cold Spring Harbor Laboratory Pr,2000:
[97]Gatz C, Lenk I. Promoters that respond to chemical inducers[J]. Trends in plant science.1998,3(9): 352-358.
[98]Zuo J, Chua N H. Chemical-inducible systems for regulated expression of plant genes[J]. Current Opinion in Biotechnology.2000,11(2):146-151.
[99]Weinmann P, Gossen M, Hillen W, et al. A chimeric transactivator allows tetracycline-responsive gene expression in whole plants[J]. The Plant Journal.2003,5(4):559-569.
[100]Zhu Q, Dabi T, Lamb C. TATA box and initiator functions in the accurate transcription of a plant minimal promoter in vitro[J]. The Plant Cell Online.1995,7(10):1681-1689.
[101]Neve R L, West R W, Rodriguez R L. Eukaryotic DNA fragments which act as promoters for a plasmid gene[J]. Natur.1979,277(5694):324-325.
[102]Sidhu R S, Mathewes S, Bollon A P. Selection of secretory protein-encoding genes by fusion with PHO5 in Saccharomyces cerevisiae[J]. Gene.1991,107(1):111-118.
[103]李姗姗,迟彦,李凌飞等.启动子克隆方法研究进展[J].中国生物工程杂志.2005,25(7)：9-16.
[104]Vida T A, Graham T R, Emr S D. In vitro reconstitution of intercompartmental protein transport to the yeast vacuole[J]. Journal of Cell Biology.1990,111(6):2871-2884.
[105]Friedrich G, Soriano P. Promoter traps in embryonic stem cells:a genetic screen to identify and mutate developmental genes in mice[J]. Genes Dev.1991,5(9):1513-1523.
[106]Lindsey K, Wei W, Clarke M C, et al. Tagging genomic sequences that direct transgene expression by activation of a promoter trap in plants[J]. Transgenic research.1993,2(1):33-47.
[107]王爱民,陈石,燕一等.Ac/Ds (GUS)介导的水稻启动子捕获系统的建立[J].植物生理与分子生物学学报.2005,31(6)：575-580.
[108]方华舟,涂知明.水稻花粉组织特异性启动子捕获实验[J].生物学杂志.2007,24(5)：37-39.
[1091李竹红,刘德培,梁植权.改进的反向PCR技术克隆转移基因的旁侧序列[J].生物化学与生物物理进展.1999,26(6)：600-602.
[110]Triglia T, Peterson M, Kemp D. A procedure for in vitro amplification of DNA segments that lie outside the boundaries of known sequences[J]. Nucleic Acids Research.1988,16(16):8186.
[111]Jones D H, Winistorfer S C. Recombinant circle PCR and recombination PCR for site-specific mutagenesis without PCR product purification[J]. BioTechniques.1992,12(4):528-535.
[112]Jones D H, Winistorfer S C. Amplification of 4-9-kb human genomic DNA flanking a known site using a panhandle PCR variant[J]. BioTechniques.1997,23(1):132-138.
[113]Prashar Y, Weissman S M. Analysis of differential gene expression by display of 3'end restriction fragments of cDNAs[J]. Proceedings of the National Academy of Sciences.1996,93(2):659-663.
[114]Shyamala V, Ames G F. Amplification of bacterial genomic DNA by the polymerase chain reaction and direct sequencing after asymmetric amplification:application to the study of periplasmic permeases[J]. Journal of bacteriology.1989,171(3):1602-1608.
[115]Han J, Luhs W, Sonntag K, et al. Functional characterisation of p-ketoacyl-CoA synthase genes from Brassica napus [J]. L. Plant Mol. Biol.2001,46:229-239.
[116]王新国,张国华.用衔接头PCR克隆新的胡萝卜Ⅱ型转化酶基因启动子[J].中国生物化学与分子生物学报.2001,17(1)：61-65.
[117]Liu Y G, Whittier R F. Thermal asymmetric interlaced PCR:automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking [J]. Genomics.1995,25(3):674-681.
[118]Chen S, Jin W, Wang M, et al. Distribution and characterization of over 1000 T-DNA tags in rice genome[J]. The Plant Journal.2003,36(1):105-113.
[119]Zhang Q. Strategies for developing green super rice[J]. Proceedings of the National Academy of Sciences.2007,104(42):16402-16409.
[120]Schmid C D, Praz V, Delorenzi M, et al. The Eukaryotic Promoter Database EPD:the impact of in silico primer extension[J]. Nucleic Acids Research.2004,32:82-85.
[121]Heinemeyer T, Wingender E, Reuter I, et al. Databases on transcriptional regulation:TRANSFAC, TRRD and COMPEL[J]. Nucleic Acids Research.1998,26(1):362-367.
[122]Higo K, Ugawa Y, Iwamoto M, et al. Plant cis-acting regulatory DNA elements (PLACE) database:1999[J]. Nucleic Acids Research.1999,27(1):297-300.
[123]Rombauts S, Dehais P, Van Montagu M, et al. PlantCARE, a plant cis-acting regulatory element database[J]. Nucleic Acids Research.1999,27(1):295-296.
[124]朱廷恒,罗红丽,宋凤鸣等.水稻病程相关蛋白基因OsPR-4b启动子的克隆及缺失体构建fJl.浙江大学学报：农业与生命科学版,2005,31(1)：22-26.
[125]杨英军,周鹏.番木瓜proteinase omega基因启动子的克隆及功能初步研究[J].云南植物研究.2005,27(5)：545-551.
[126]宗成文,章镇,房经贵等.葡萄LEAFY基因启动子的克隆与序列分析[J].南京农业大学学报.2007,30(4)：20-25.
[127]Jefferson R A, Kavanagh T A, Bevan M W. GUS fusions:beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants[J]. The EMBO Journal.1987,6(13):3901-3907.
[128]Baurle I, Laux T. Regulation of WUSCHEL transcription in the stem cell niche of the Arabidopsis shoot meristem[J]. The Plant Cell Online.2005,17(8):2271-2280.
[129]Matarasso N, Schuster S, Avni A. A novel plant cysteine protease has a dual function as a regulator of 1-aminocyclopropane-l-carboxylic acid synthase gene expression[J]. The Plant Cell Online.2005,17(4):1205-1216.
[130]王新力,彭学贤.香蕉果实成熟相关基因ACO1启动子区的克隆及其功能初探[J].生物工程学报.2001,17(4)：428-431.
[131]Atkinson R G, Bolitho K M, Wright M A, et al. Apple ACC-oxidase and polygalacturonase: ripening-specific gene expression and promoter analysis in transgenic tomato[J]. Plant Molecular Biology.1998,38(3):449-460.
[132]Wang Z Y, MacRae E A, Wright M A, et al. Polygalacturonase gene expression in kiwifruit: relationship to fruit softening and ethylene production[J]. Plant Molecular Biology.2000,42(2): 317-328.
[133]袁正强,吴家和.三个韧皮部特异性启动子在转基因烟草中表达的比较研究[J].农业生物技术学报.2002,10(1)：6-9.
[134]Melegari M, Scaglioni P P, Wands J R. Cloning and characterization of a novel hepatitis B virus x binding protein that inhibits viral replication[J]. Journal of virology.1998,72(3):1737-1743.
[135]Liao Y, Zou H F, Wei W, et al. Soybean GmbZIP44, GmbZIP62 and GmbZIP78 genes function as negative regulator of ABA signaling and confer salt and freezing tolerance in transgenic Arabidopsis[J]. Planta.2008,228(2):225-240.
[136]Wang S. Identification of novel pathogen-responsive cis-elements and their binding proteins in the promoter of OsWRKY13, a gene regulating rice disease resistance[J]. Plant, Cell and Environment.2007,31:86-96.
[137]Galas D J, Schmitz A. DNase I footprinting; a simple method for the detection of protein-DNA binding specificty[J]. Nucleic Acids Res.1978,5:3157-3170.
[138]Ackers G, Johnson A, Shea M. Quantitative model for gene regulation by lambda phage repressor[J]. Proc Natl Acad Sci USA.1982,79(4):1129-1136.
[139]Nakamura I, Koike K. Identification of a binding protein to the X gene promoter region of hepatitis B virus[J]. Virology.1992,191(2):533-540.
[140]Bouffartigues E, Buckle M, Badaut C, et al. H-NS cooperative binding to high-affinity sites in a regulatory element results in transcriptional silencing[J]. Nat Struct Mol Biol.2007,14(5): 441-448.
[141]Adams B, Cha H, Cleary J, et al. DEK binding to class Ⅱ MHC Y-box sequences is gene-and allele-specific[J]. Arthritis Res Ther. 2003,5(4):226-233.
[142]Flanigan A, Gardner J F. Interaction of the Gifsy-1 Xis protein with the Gifsy-1 attP sequence[J]. Journal of bacteriology.2007,189(17):6303-6311.
[143]Smith J V, Luo Y. Studies on molecular mechanisms of Ginkgo biloba extract[J]. Applied microbiology and biotechnology.2004,64(4):465-472.
[144]Mehdy M, Lamb C. Chalcone isomerase cDNA cloning and mRNA induction by fungal elicitor, wounding and infection[J]. The EMBO Journal.1987,6(6):1527.
[145]van Tunen A J, Koes R E, Spelt C E, et al. Cloning of the two chalcone flavanone isomerase genes from Petunia hybrida:coordinate, light-regulated and differential expression of flavonoid genes[J]. EMBO J.1988,7(5):1257-1263.
[146]Blyden E R, Doerner P W, Lamb C J, et al. Sequence analysis of a chalcone isomerase cDNA of Phaseolus vulgaris L[J]. Plant Mol Biol.1991,16(1):167-169.
[147]Grotewold E, Peterson T. Isolation and characterization of a maize gene encoding chalcone flavonone isomerase[J]. Mol Gen Genet.1994,242(1):1-8.
[148]Wood A J, Davies E. A cDNA Encoding Chalcone Isomerase from Aged Pea Epicotyls[J]. Plant Physiology.1994,104(4):1465-1466.
[149]McKhann H I, Hirsch A M. Isolation of chalcone synthase and chalcone isomerase cDNAs from alfalfa (Medicago sativa L.):highest transcript levels occur in young roots and root tips[J]. Plant Mol Biol.1994,24(5):767-777.
[150]Li F, Jin Z, Qu W, et al. Cloning of a cDNA encoding the Saussurea medusa chalcone isomerase and its expression in transgenic tobacco[J]. Plant Physiol Biochem.2006,44(7-9):455-461.
[151]Gensheimer M, Mushegian A. Chalcone isomerase family and fold:No longer unique to plants [J]. Protein Sci.2004,13(2):540-544.
[152]Druka A, Kudrna D, Rostoks N, et al. Chalcone isomerase gene from rice (Oryza sativa) and barley (Hordeum vulgare):physical, genetic and mutation mapping[J]. Gene.2003,302(1-2): 171-178.
[153]Shimada N, Aoki T, Sato S, et al. A Cluster of genes encodes the two types of chalcone isomerase involved in the biosynthesis of general flavonoids and legume-specific 5-deoxy(iso)flavonoids in Lotus japonicus[J]. Plant Physiology.2003,131(3):941-951.
[154]McClean P E, Lee R K. Genetic architecture of chalcone isomerase non-coding regions in common bean (Phaseolus vulgaris L.)[J]. Genome.2007,50(2):203-214.
[155]Kuittinen H, Aguade M. Nucleotide variation at the chalcone isomerase locus in Arabidopsis thaliana[J]. Genetics.2000,155(2):863-872.
[156]Van Tunen A, Mur L, Brouns G, et al. Pollen-and anther-specific chi promoters from petunia: tandem promoter regulation of the chiA gene[J]. The Plant Cell Online.1990,2(5):393-401.
[157]程水源,陈昆松,杜何为等.银杏RNA的提取[J].果树学报.2005,22(4)：428-429.
[158]Sambrook J, Russell D W. Molecular cloning:a laboratory manual[M]. CSHL press,2001:
[159]魏春红,李毅.现代分子生物学实验[M].北京：高等教育出版社,2006：135-137.
[160]Jansson S, Meyer-Gauen G, Cerff R, et al. Nucleotide distribution in gymnosperm nuclear sequences suggests a model for GC-content change in land-plant nuclear genomes[J]. Journal of molecular evolution.1994,39(1):34-46.
[161]Jez J M, Noel J P. Reaction mechanism of chalcone isomerase:pH dependence, diffusion control, and product binding differences [J]. J. Biol. Chem.2002,277(2):1361-1369.
[162]Fouche S D, Dubery I A. Chalcone isomerase from Citrus sinensis:purification and characterization[J]. Phytochemistry.1994,37(1):127-132.
[163]Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding[J]. Analytical biochemistry.1976,72(1-2): 248-254.
[164]Moustafa E, Wong E. Purification and properties of chalcone-flavanone isomerase from soya bean seed[J]. Phytochemistry.1967,6(5):625-632.
[165]程水源,王燕.银杏叶黄酮含量变化及分布规律的研究[J].园艺学报.2001,28(4)：353-355.
[166]Lutcke H A, Chow K C, Mickel F S, et al. Selection of AUG initiation codons differs in plants and animals[J]. EMBO J.1987,6(1):43-48.
[167]Folter S, Angenent G. trans meets cis in MADS science[J]. Trends in plant science.2006,11(5): 224-231.
[168]Yang T, Poovaiah B. A calmodulin-binding/CGCG box DNA-binding protein family involved in multiple signaling pathways in plants[J]. Journal of Biological Chemistry.2002,277 (47): 45049-45058.
[169]Piechulla B, Merforth N, Rudolph B. Identification of tomato Lhc promoter regions necessary for circadian expression[J]. Plant Molecular Biology.1998,38(4):655-662.
[170]Chen C, Chen Z. Potentiation of developmentally regulated plant defense response by AtWRKY18, a pathogen-induced Arabidopsis transcription factor[J]. Plant Physiology.2002, 129(2):706-716.
[171]Schunmann P, Richardson A, Vickers C, et al. Promoter analysis of the barley Phtl; 1 phosphate transporter gene identifies regions controlling root expression and responsiveness to phosphate deprivation[J]. Plant Physiology.2004,136(4):4205-4214.
[172]Dunn M, White A, Vural S, et al. Identification of promoter elements in a low-temperature-responsive gene (blt4.9) from barley(Hordeum vulgare L.)[J]. Plant Molecular Biology.1998, 38(4):551-564.
[173]Hartmann U, Sagasser M, Mehrtens F, et al. Differential combinatorial interactions of cis-acting elements recognized by R2R3-MYB, BZIP, and BHLH factors control light-responsive and tissue-specific activation of phenylpropanoid biosynthesis genes[J]. Plant Mol Biol.2005,57(2): 155-171.
[174]Bate N, Twell D. Functional architecture of a late pollen promoter:pollen-specific transcription is developmentally regulated by multiple stage-specific and co-dependent activator elements[J]. Plant Molecular Biology.1998,37(5):859-869.
[175]Yoshioka S, Taniguchi F, Miura K, et al. The novel Myb transcription factor LCR1 regulates the CO2-responsive gene Cahl, encoding a periplasmic carbonic anhydrase in Chlamydomonas reinhardtii[J]. Plant Cell.2004,16(6):1466-1477.
[176]Solano R, Nieto C, Avila J, et al. Dual DNA binding specificity of a petal epidermis-specific MYB transcription factor (MYB. Ph3) from Petunia hybrida[J]. The EMBO Journal.1995,14(8): 1773-1784.
[177]Lam E, Chua N. ASF-2:a factor that binds to the cauliflower mosaic virus 35S promoter and a conserved GATA motif in Cab promoters[J]. The Plant Cell Online.1989,1(12):1147-1156.
[178]Lopez-Molina L, Chua N H. A null mutation in a bZIP factor confers ABA-insensitivity in Arabidopsis thaliana[J]. Plant and Cell Physiology.2000,41(5):541-547.
[179]Ogawa M, Hanada A, Yamauchi Y, et al. Gibberellin biosynthesis and response during Arabidopsis seed germination[J]. The Plant Cell Online.2003,15(7):1591-1604.
[180]Boyle B, Brisson N. Repression of the defense gene PR-10a by the single-stranded DNA binding protein SEBF[J]. The Plant Cell Online.2001,13(11):2525-2537.
[181]Yamagata H, Yonesu K, Hirata A, et al. TGTCACA Motif Is a Novel cis-Regulatory Enhancer Element Involved in Fruit-specific Expression of thecucumisin Gene[J]. Journal of Biological Chemistry.2002,277(13):11582-11590.
[182]Zhou D. Regulatory mechanism of plant gene transcription by GT-elements and GT-factors[J]. Trends in plant science.1999,4(6):210-214.
[183]Park H, Kim M, Kang Y, et al. Pathogen-and NaCl-induced expression of the SCaM-4 promoter is mediated in part by a GT-1 box that interacts with a GT-1-like transcription factor[J]. Plant Physiology.2004,135(4):2150-2161.
[184]Rawat R, Xu Z-F, Yao K-M, et al. Identification of cis-elements for ethylene and circadian regulation of the Solanum melongena gene encoding cysteine proteinase[J]. Plant Mol Biol.2005, 57(5):629-643.
[185]Itzhaki H, Maxson J M, Woodson W R. An ethylene-responsive enhancer element is involved in the senescence-related expression of the carnation glutathione-S-transferase (GST1) gene[J]. PNAS.1994,91(19):8925-8929.
[186]Montgomery J, Goldman S, Deikman J, et al. Identification of an ethylene-responsive region in the promoter of a fruit ripening gene[J]. PNAS.1993,90(13):5939-5943.
[187]Jez J M, Bowman M E, Dixon R A, et al. Structure and mechanism of the evolutionarily unique plant enzyme chalcone isomerase[J]. Nat Struct Biol.2000,7(9):786-791.
[188]Bednar R, Hadcock J. Purification and characterization of chalcone isomerase from soybeans[J]. Journal of Biological Chemistry.1988,263(20):9582-9588.
[189]Dixon R, Richard Blyden E, Robbins M, et al. Comparative biochemistry of chalcone isomerases[J]. Phytochemistry.1988,27(9):2801-2808.
[190]Shirley B, Kubasek W, Storz G, et al. Analysis of Arabidopsis mutants deficient in flavonoid biosynthesis[J]. The Plant Journal.1995,8(5):659-671.
[191]Xu F, Cheng S Y, Cheng S H, et al. Time course of expression of chalcone synthase gene in Ginkgo biloba[J]. Journal of Plant Physiology and Molecular Biology.2007,33(4):309-317.
[192]Li J, Ou-Lee T, Raba R, et al. Arabidopsis flavonoid mutants are hypersensitive to UV-B irradiation[J]. The Plant Cell Online.1993,5(2):171-179.
[193]Bieza K, Lois R. An Arabidopsis mutant tolerant to lethal ultraviolet-B levels shows constitutively elevated accumulation of flavonoids and other phenolics[J]. Plant Physiology.2001,126(3): 1105-1115.
[194]Kush A, Goyvaerts E, Chye M L, et al. Laticifer-specific gene expression in Hevea brasiliensis (rubber tree)[J]. Proceedings of the National Academy of Sciences.1990,87(5):1787-1790.
[195]Ardi R, Kobiler I, Jacoby B, et al. Involvement of epicatechin biosynthesis in the activation of the mechanism of resistance of avocado fruits to Colletotrichum gloeosporioides[J]. Physiological and Molecular Plant Pathology.1998,53(5-6):269-285.
[196]Abe H, Yamaguchi-Shinozaki K, Urao T, et al. Role of Arabidopsis MYC and MYB homologs in drought-and abscisic acid-regulated gene expression[J]. The Plant Cell Online.1997,9(10): 1859-1868.
[197]蔡葛平,郭燕红,姚辉等.矮壮素和赤霉素对黄芩生物量及根中黄酮类成分产量的影响[J].中国农学通报.2008,24(7)：213-217.
[198]程水源,王燕,李俊凯等.内源激素含量与银杏叶中类黄酮含量的关系[J].林业科学.2004,40(6)：45-49.
[199]Von Wettstein D, Gough S, Kannangara C. Chlorophyll biosynthesis[J]. The Plant Cell.1995,7(7): 1039-1057.
[200]汪良驹,王中华,李志强等.5-氨基乙酰丙酸促进苹果果实着色的效应[J].果树学报.2004,21(006)：512-515.
[201]程水源,王燕,李俊凯等.银杏叶片色素含量与黄酮含量关系的研究[J].林业科学.2001,37(5)：31-34.
[202]Brugliera F, Barri-Rewell G, Holton T, et al. Isolation and characterization of a flavonoid 3'-hydroxylase cDNA clone corresponding to the Htl locus of Petunia hybrida[J]. The Plant Journal.2002,19(4):441-451.
[203]Seitz C, Eder C, Deiml B, et al. Cloning, functional identification and sequence analysis of flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase cDNAs reveals independent evolution of flavonoid 3',5'-hydroxylase in the Asteraceae family[J]. Plant Molecular Biology.2006,61(3): 365-381.
[204]Tanaka Y. Flower colour and cytochromes P450[J]. Phytochemistry Reviews.2006,5(2): 283-291.
[205]Nakatsuka T, Nishihara M, Mishiba K, et al. Heterologous expression of two gentian cytochrome P450 genes can modulate the intensity of flower pigmentation in transgenic tobacco plants[J]. Molecular Breeding.2006,17(2):91-99.
[206]Jeong S, Goto-Yamamoto N, Hashizume K, et al. Expression of the flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase genes and flavonoid composition in grape (Vitis vinifera)[J]. Plant
Science.2006,170(1):61-69.
[207]Kaplan B, Davydov O, Knight H, et al. Rapid transcriptome changes induced by cytosolic Ca2+ transients reveal ABRE-related sequences as Ca2+-responsive cis elements in Arabidopsis[J]. Plant Cell.2006,18(10):2733-2748.
[208]Luo H, Song F, Goodman R M, et al. Up-regulation of OsBIHD1, a rice gene encoding BELL homeodomain transcriptional factor, in disease resistance responses[J]. Plant Biology.2005,7(5): 459-468.
[209]Abe H, Urao T, Ito T, et al. Arabidopsis AtMYC2 (bHLH) and AtMYB2 (MYB) function as transcriptional activators in abscisic acid signaling[J]. The Plant Cell Online.2003,15(1):63-67.
[210]Fujiwara T, Beachy R N. Tissue-specific and temporal regulation of a β-conglycinin gene:roles of the RY repeat and other cis-acting elements[J]. Plant Molecular Biology.1994,24(2):261-272.
[211]Plesch G, Ehrhardt T, Mueller-Roeber B. Involvement of TAAAG elements suggests a role for Dof transcription factors in guard cell-specific gene expression[J]. The Plant Journal.2002,28(4): 455-464.
[212]Kram B, Xu W, Carter C. Uncovering the Arabidopsis thaliana nectary transcriptome: investigation of differential gene expression in floral nectariferous tissues[J]. BMC plant biology. 2009,9(1):92.
[213]Tang W, Perry S. Binding site selection for the plant MADS domain protein AGL15[J]. Journal of Biological Chemistry.2003,278(30):28154.
[214]Ellerstr m M, St lberg K, Ezcurra I, et al. Functional dissection of a napin gene promoter: identification of promoter elements required for embryo and endosperm-specific transcription[J]. Plant Molecular Biology.1996,32(6):1019-1027.
[215]Fusada N, Masuda T, Kuroda H, et al. Identification of a novel cis-element exhibiting cytokinin-dependent protein binding in vitro in the 5'-region of NADPH-protochlorophyllide oxidoreductase gene in cucumber[J]. Plant Molecular Biology.2005,59(4):631-645.
[216]Kim S, Chung H, Thomas T. Isolation of a novel class of bZIP transcription factors that interact with ABA-responsive and embryo-specification elements in the Dc3 promoter using a modified yeast one-hybrid system[J]. The Plant Journal.2002,11(6):1237-1251.
[217]Eulgem T, Rushton P, Robatzek S, et al. The WRKY superfamily of plant transcription factors[J]. Trends in plant science.2000,5(5):199-205.
[218]Baumann K, De Paolis A, Costantino P, et al. The DNA binding site of the Dof protein NtBBF1 is essential for tissue-specific and auxin-regulated expression of the rolB oncogene in plants [J]. The Plant Cell Online.1999,11(3):323-334.
[219]Yamazaki S, Sato K, Suhara K, et al. Importance of the proline-rich region following signal-anchor sequence in the formation of correct conformation of microsomal cytochrome P450s[J]. Journal of biochemistry.1993,114(5):652-657.
[220]Murakami K, Mihara K, Omura T. The transmembrane region of microsomal cytochrome P450 identified as the endoplasmic reticulum retention signal[J]. Journal of biochemistry.1994,116(1): 164-173.
[221]Chapple C. Molecular-genetic analysis of plant cytochrome P450-dependent monooxygenases[J]. Annual Review of Plant Biology.1998,49(1):311-343.
[222]许志茹,崔国新,李春雷等.芜菁的类黄酮3'羟化酶基因克隆和UV-A诱导表达特性[J].植物生理学通讯.2008,44(5)：931-935.
[223]Xu B, Li J, Zhang X, et al. Cloning and molecular characterization of a functional flavonoid 3'-hydroxylase gene from Brassica napus[J]. Journal of plant physiology.2007,164(3):350-363.
[224]Schoenbohm C, Martens S, Eder C, et al. Identification of the Arabidopsis thaliana flavonoid 3'-hydroxylase gene and functional expression of the encoded P450 enzyme[J]. Biological Chemistry.2000,381(8):749-754.
[225]Xu F, Cheng H, Cai R, et al. Molecular cloning and function analysis of an anthocyanidin synthase gene from Ginkgo biloba, and its expression in abiotic stress responses[J]. Mol. Cell. 2008,26(6):536-547.
[226]Kreuzaler F, Ragg H, Fautz E, et al. UV-induction of chalcone synthase mRNA in cell suspension cultures of Petroselinum hortense[J]. Proceedings of the National Academy of Sciences of the United States of America.1983,80(9):2591.
[227]Koes R, Quattrocchio F, Mol J. The flavonoid biosynthetic pathway in plants:function and evolution[J]. BioEssays.2005,16(2):123-132.
[228]Deikman J, Hammer P E. Induction of anthocyanin accumulation by cytokinins in Arabidopsis thaliana[J]. Plant Physiology.1995,108(1):47-57.
[229]Herrmann K M, Weaver L M. The shikimate pathway[J]. Annual Review of Plant Physiology and Plant Molecular Biology.1999,50(1):473-503.
[230]Kumar A, Ellis B E. The phenylalanine ammonia-lyase gene family in raspberry. Structure, expression, and evolution[J]. Plant Physiology.2001,127(1):230-239.
[231]Hermann K M. The shikimate pathway:early steps in the biosynthesis of aromatic compounds[J]. Plant Cell.1995,7(3):907-919.
[232]Xujun W, Xiaoli W, Xugang L, et al. Isolation of rice EPSP synthase cDNA and its sequence analysis and copy number determination[J]. Acta Botanica Sinica.2002,44(2):188-192.
[233]Klee H J, Muskopf Y M, Gasser C S. Cloning of an Arabidopsis thaliana gene encoding 5-enolpyruvylshikimate-3-phosphate synthase:sequence analysis and manipulation to obtain glyphosate-tolerant plants[J]. Molecular and General Genetics.1987,210(3):437-442.
[234]刘东军,张锐,郭三堆等.棉花品系Y18在草甘膦胁迫下的epsps基因表达分析研究[J].中国生物工程杂志.2008,28(10)：55-59.
[235]蒋向,戴雄泽,李育强等.薤白EPSPs基因在不同组织表达的半定量分析[J].湖南农业大学学报(自然科学版).2007,33(5)：542-545.
[236]Gong Y, Liao Z, Chen M, et al. Characterization of 5-enolpyruvylshikimate 3-phosphate synthase gene from Camptotheca acuminata[J]. Biologia Plantarum.2006,50(4):542-550.
[237]Papanikou E, Brotherton J E, Widholm J M. Length of time in tissue culture can affect the selected glyphosate resistance mechanism[J]. Planta.2004,218(4):589-598.
[238]Stallings W C, Abdel-Meguid S S, Lim L W, et al. Structure and topological symmetry of the glyphosate target 5-enolpyruvylshikimate-3-phosphate synthase:a distinctive protein fold[J]. Proceedings of the National Academy of Sciences.1991,88(11):5046-5050.
[239]Shirley-Winkel B. It takes a garden, How work on diverse plant species ha contributed to an understanding of flavonoid metabolism[J]. Plant Physiology.2001,127(2316):1399-1404.
[240]van der Krol A R, van Poecke R M P, Vorst O F J, et al. Developmental and wound-, cold-, desiccation-, ultraviolet-B-stress-induced modulations in the expression of the petunia zinc finger transcription factor gene ZPT2-2[J]. Plant Physiology.1999,121(4):1153-1162.
[241]Benfey P N, Takatsuji H, Ren L, et al. Sequence requirements of the 5-enolpyruvylshikimate-3-phosphate synthase 5 [prime]-upstream region for tissue-specific expression in flowers and seedlings[J]. Plant Cell.1990,2(9):849-856.
[242]魏建华,宋艳茹.木质素生物合成途径及调控的研究进展[J].植物学报：英文版.2001,43(008)：771-779.
[243]Lacombe E, Hawkins S, Van Doorsselaere J, et al. Cinnamoyl CoA reductase, the first committed enzyme of the lignin branch biosynthetic pathway:cloning, expression and phylogenetic relationships[J]. The Plant Journal.2003,11(3):429-441.
[244]Piquemal J, Lapierre C, Myton K, et al. Down-regulation of cinnamoyl-CoA reductase induces significant changes of lignin profiles in transgenic tobacco plants[J]. The Plant Journal.2002, 13(1):71-83.
[245]Jones L, Ennos A R, Turner S R. Cloning and characterization of irregular xylem4 (irx4):a severely lignin-deficient mutant of Arabidopsis[J]. The Plant Journal.2001,26(2):205-216.
[246]李波,梁颖,柴友荣.植物肉桂酰辅酶A还原酶(CCR)基因的研究进展[J].分子植物育种.2006,4(3)：55-65.
[247]Sarni F, Grand C, Boudet A. Purification and properties of cinnamoyl-CoA reductase and cinnamyl alcohol dehydrogenase from poplar stems (Populus×euramericana)[J]. European Journal of Biochemistry.2005,139(2):259-265.
[248]Goffner D, Campbell M, Campargue C, et al. Purification and characterization of cinnamoyl-coenzyme A:NADP oxidoreductase in Eucalyptus gunnii[J]. Plant Physiology.1994, 106(2):625-632.
[249]Luderitz T, Grisebach H. Enzymic synthesis of lignin precursors comparison of cinnamoyl-CoA reductase and cinnamyl alcohol:NADP+ dehydrogenase from spruce (Picea abies L.) and soybean (Glycine max L.)[J]. European Journal of Biochemistry.2005, 119(1):115-124.
[250]McInnes R, Lidgett A, Lynch D, et al. Isolation and characterization of a cinnamoyl-CoA reductase gene from perennial ryegrass (Lolium perenne)[J]. Journal of plant physiology.2002, 159(4):415-422.
[251]Larsen K. Cloning and characterization of a ryegrass (Lolium perenne) gene encoding cinnamoyl-CoA reductase (CCR) [J]. Plant Science.2004,166(3):569-581.
[252]Lewis N, Davin L. Evolution of lignan and neolignan biochemical pathways:American chemical society [J]. Isopentenoids and other natural products:evolution and function.1994,202-246.
[253]Li L, Cheng X, Lu S, et al. Clarification of cinnamoyl co-enzyme A reductase catalysis in monolignol biosynthesis of aspen[J]. Plant and Cell Physiology.2005,46(7):1073-1082.
[254]Lauvergeat V, Lacomme C, Lacombe E, et al. Two cinnamoyl-CoA reductase (CCR) genes from Arabidopsis thaliana are differentially expressed during development and in response to infection with pathogenic bacteria [J]. Phytochemistry.2001,57(7):1187-1195.
[255]Pichon M, Courbou I, Beckert M, et al. Cloning and characterization of two maize cDNAs encoding cinnamoyl-CoA reductase (CCR) and differential expression of the corresponding genes[J]. Plant Molecular Biology.1998,38(4):671-676.
[256]Wengenmayer H, Ebel J, Grisebach H. Purification and properties of a cinnamoyl CoA:NADPH reductase from cell suspension cultures of soybean (Glycine max) [J]. European Journal of Biochemistry.65(2):529-536.
[257]Lacombe E, Van Doorsselaere J, Boerjan W, et al. Characterization of cis-elements required for vascular expression of the cinnamoyl CoA reductase gene and for protein-DNA complex formation[J]. The Plant Journal.2000,23(5):663-676.
[258]Selman-Housein G, Lopez M, Hernandez D, et al. Molecular cloning of cDNAs coding for three sugarcane enzymes involved in lignification[J]. Plant Sci.1999,143(2):163-171.
[259]Raven J. The evolution of vascular land plants in relation to supracellular transport processes[J]. Advances in botanical research.1977,5(1):153-219.
[260]van Beek T. Chemical analysis of Ginkgo biloba leaves and extracts[J]. Journal of Chromatography A.2002,967(1):21-55.
[261]Heller W, Hahlbrock K. Highly purified" flavanone synthase" from parsley catalyzes the formation of naringenin chalcone[J]. Archives of biochemistry and biophysics.1980,200(2): 617-619.
[262]许锋.银杏查尔酮合成酶基因和苯丙氨酸解氨酶基因的克隆及表达[D].武汉：华中农业大学,2005
[263]Dynan W. Modularity in promoters and enhancers[J]. Cell.1989,58(1):1-4.
[264]Loake G J, Faktor O, Lamb C J, et al. Combination of H-box [CCTACC (N) 7CT] and G-box (CACGTG) cis elements is necessary for feed-forward stimulation of a chalcone synthase promoter by the phenylpropanoid-pathway intermediate p-coumaric acid[J]. PNAS.1992,89(19): 9230-9234.
[265]Schulze-Lefert P, Becker-Andre M, Schulz W, et al. Functional architecture of the light-responsive chalcone synthase promoter from parsley[J]. Plant Cell.1989,1(7):707-714.
[266]Hartmann U, Valentine W J, Christie J M, et al. Identification of UV/blue light-response elements in the Arabidopsis thaliana chalcone synthase promoter using a homologous protoplast transient expression system[J]. Plant Molecular Biology.1998,36(5):741-754.
[267]Lawton M A, Dean S M, Dron M, et al. Silencer region of a chalcone synthase promoter contains multiple binding sites for a factor, SBF-1, closely related to GT-1[J]. Plant Molecular Biology. 1991,16(2):235-249.
[268]van der Meer I M, Brouwer M, Spelt C E, et al. The TACPyAT repeats in the chalcone synthase promoter of Petunia hybrida act as a dominant negative cis-acting module in the control of organ-specific expression[J]. The Plant Journal.1992,2(4):525-535.
[269]Wang J, Letham D S, Cornish E, et al. Studies of cytokinin action and metabolism using tobacco plants expressing either the ipt or the GUS gene controlled by a chalcone synthase Promoter. I. Developmental Features of the Transgenic Plants[J]. Functional Plant Biology.1997,24(5): 661-672.
[270]Faktor O, Kooter J M, Dixon R A, et al. Functional dissection of a bean chalcone synthase gene promoter in transgenic tobacco plants reveals sequence motifs essential for floral expression[J]. Plant Molecular Biology.1996,32(5):849-859.
[271]许锋,程水源,王燕等.TAIL-PCR方法快速克隆银杏查尔酮合成酶基因及序列分析[J].果树学报.2007,24(2)：237-243.
[272]Jefferson R A. Assaying chimeric genes in plants:the GUS gene fusion system[J]. Plant Molecular Biology Reporter.1987,5(4):387-405.
[273]Thum K E, Kim M, Morishige D T, et al. Analysis of barley chloroplast psbD light-responsive promoter elements in transplastomic tobacco[J]. Plant Molecular Biology.2001,47(3):353-366.
[274]Block A, Dangl J L, Hahlbrock K, et al. Functional borders, genetic fine structure, and distance requirements of cis elements mediating light responsiveness of the parsley chalcone synthase promoter[J]. PNAS.1990,87(14):5387-5391.
[275]Svensson J T, Crosatti C, Campoli C, et al. Transcriptome analysis of cold acclimation in barley Albina and Xantha mutants[J]. Plant Physiology.2006,141(1):257-270.
[276]Simpson S D, Nakashima K, Narusaka Y, et al. Two different novel cis-acting elements of erd 1, a clpA homologous Arabidopsis gene function in induction by dehydration stress and dark-induced senescence[J]. The Plant Journal.2003,33(2):259-270.
[277]Rushmore T H, Morton M R, Pickett C B. The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity[J]. Journal of Biological Chemistry.1991,266(18):11632-11639.
[278]Quinn J M, Eriksson M, Moseley J L, et al. Oxygen deficiency responsive gene expression in Chlamydomonas reinhardtii through a copper-sensing signal transduction pathway[J]. Plant Physiology.2002,128(2):463-471.
[279]Quinn J M, Barraco P, Eriksson M, et al. Coordinate copper-and oxygen-responsive Cyc6 and Cpxl expression in Chlamydomonas is mediated by the same element[J]. Journal of Biological Chemistry.2000,275(9):6080-6089.
[280]Saha D, Prasad A, Sujatha T, et al. In silico analysis of the Lateral Organ Junction (loj) gene and promoter of Arabidopsis thaliana[J]. In Silico Biology.2007,7(1):7-19.
[281]Urao T, Yamaguchi-Shinozaki K, Urao S, et al. An Arabidopsis myb homolog is induced by dehydration stress and its gene product binds to the conserved MYB recognition sequence[J]. Plant Cell.1993,5(11):1529-1539.
[282]Zhu Y, Cai X L, Wang Z Y, et al. An interaction between a MYC protein and an EREBP protein is involved in transcriptional regulation of the rice Wx gene[J]. Journal of Biological Chemistry. 2003,278(48):47803-47811.
[283]Kucho K, Yoshioka S, Taniguchi F, et al. Cis-acting elements and DNA-binding proteins involved in CO2-responsive transcriptional activation of Cahl encoding a periplasmic carbonic anhydrase in Chlamydomonas reinhardtii[J]. Plant Physiology.2003,133(2):783-793.
[284]Hamilton D A, Schwarz Y H, Mascarenhas J P. A monocot pollen-specific promoter contains separable pollen-specific and quantitative elements[J]. Plant Molecular Biology.1998,38(4): 663-669.
[285]Inukai Y, Sakamoto T, Ueguchi-Tanaka M, et al. Crown rootlessl, which is essential for crown root formation in rice, is a target of an AUXIN RESPONSE FACTOR in auxin signaling[J]. Plant Cell.2005,17(5):1387-1396.
[286]Harper R M, Stowe-Evans E L, Luesse D R,et al. The NPH4 locus encodes the auxin response factor ARF7, a conditional regulator of differential growth in aerial Arabidopsis tissue[J]. Plant Cell.2000,12(5):757-770.
[287]Liu Z B, Ulmasov T, Shi X, et al. Soybean GH3 promoter contains multiple auxin-inducible elements[J]. Plant Cell.1994,6(5):645-657.
[288]Finkelstein R R, Lynch T J. The Arabidopsis abscisic acid response gene ABI5 encodes a basic leucine zipper transcription factor[J]. Plant Cell.2000,12(4):599-609.
[289]Zhang Z L, Xie Z, Zou X, et al. A rice WRKY gene encodes a transcriptional repressor of the gibberellin signaling pathway in aleurone cells[J]. Plant Physiology.2004,134(4):1500-1513.
[290]Xie Z, Zhang Z L, Zou X, et al. Annotations and functional analyses of the rice WRKY gene superfamily reveal positive and negative regulators of abscisic acid signaling in aleurone cells[J]. Plant Physiology.2005,137(1):176-189.
[291]Nishiuchi T, Shinshi H, Suzuki K. Rapid and transient activation of transcription of the ERF3 gene by wounding in tobacco leaves:possible involvement of NtWRKYs and autorepression[J]. Journal of Biological Chemistry.2004,279(53):55355-55361.
[292]Hinderer W, Petersen M, Seitz H U. Inhibition of flavonoid biosynthesis by gibberellic acid in cell suspension cultures of Daucus carota L[J]. Planta.1984,160(6):544-549.
[293]Finkler A, Kaplan B, Fromm H. Ca2+-responsive cis-elements in plants[J]. Plant Signaling & Behavior.2007,2(1):17-19.
[294]Yanagisawa S, Sheen J. Involvement of maize Dof zinc finger proteins in tissue-specific and light-regulated gene expression[J]. Plant Cell.1998,10(1):75-90.
[295]Schmid J, Doerner P, Clouse S, et al. Developmental and environmental regulation of a bean chalcone synthase promoter in transgenic tobacco[J]. Plant Cell.1990,2(7):619-631.
[296]Kubasek W, Shirley B, McKillop A, et al. Regulation of flavonoid biosynthetic genes in germinating Arabidopsis seedlings[J]. Plant Cell.1992,4(10):1229-1236.
[2]程水源,顾曼如,束怀瑞.银杏叶黄酮研究进展[J].林业科学.2000,36(6)：110-115.
[3]毕殉,卢嘉文,蔡东联.银杏及叶中黄酮类化合物生理功效的研究进展[J].武警医学.2004,15(6)：458-459.
[4]李勇,刘新民.银杏及叶中黄酮类化合物生理功效的研究进展[J].食品科技.2001,(5)：72-73.
[5]Cook N, Samman S. Flavonoids-chemistry, metabolism, cardioprotective effects, and dietary sources[J]. The Journal of nutritional biochemistry.1996,7(2):66-76.
[6]Winkel-Shirley B. Biosynthesis of flavonoids and effects of stress [J]. Current opinion in plant biology.2002,5(3):218-223.
[7]Tahara S, Ibrahim R K. Prenylated isoflavonoids-an update[J]. Phytochemistry.1995,38(5): 1073-1094.
[8]Dooner H, Robbins T, Jorgensen R. Genetic and developmental control of anthocyanin biosynthesis[J]. Annual Review of Genetics.1991,25(1):173-199.
[9]Koes R, Quattrocchio F, Mol J. The flavonoid biosynthetic pathway in plants:function and evolution[J]. BioEssays.1994,16(2):123-132.
[10]Dixon R, Paiva N. Stress-induced phenylpropanoid metabolism[J]. The Plant Cell.1995,7(7): 1085.
[11]Schijlen E, Ric de Vos C, van Tunen A, et al. Modification of flavonoid biosynthesis in crop plants[J]. Phytochemistry.2004,65(19):2631-2648.
[12]梁立兴.中国当代银杏大全[M].1993：1-41.
[13]程水源.银杏叶化学成分与药用研究进展[J].中草药.1998,(29)：16-18.
[14]邵继平,王伯初.银杏叶提取物药用价值的研究进展[J].重庆大学学报：自然科学版.2003,26(1)：130-134.
[15]古勇,李安明.类黄酮生物活性的研究进展[J].应用与环境生物学报.2006,12(2)：283-286.
[16]Hasler A, Gross G A, Meier B, et al. Complex flavonol glycosides from the leaves of Ginkgo biloba[J]. Phytochemistry.1992,31(4):1391.
[17]夏晓晖,张宇,郗砚彬等.银杏叶化学成分研究进展[J].中国实验方剂学杂志.2009,15(9)：100-104.
[18]唐于平,王颖.银杏叶中的黄酮醇苷类成分[J].药学学报.2000,35(5)：363-366.
[19]唐于平,楼凤昌.银杏叶中黄酮类成分的研究[J].中国药学杂志.2001,36(4)：231-233.
[20]Dixon R A, Steele C L. Flavonoids and isoflavonoids-a gold mine for metabolic engineering[J]. Trends in plant science.1999,4(10):394-400.
[21]Holton T A, Cornish E C. Genetics and biochemistry of anthocyanin biosynthesis[J]. The Plant Cell. 1995,7(7):1071.
[22]Tsai C J, El Kayal W, Harding S A. Populus, the new model system for investigating phenylpropanoid complexity [J]. Int J Appl Sci Eng.2006,4:221-233.
[23]Pla J, Ville A, Pacheco H. Biogenesis of plant pigments.1. Comparative study of the incorporation of 1,2-14C shikimic and 3-14C trans-cinnamic acids in two anthocyanic pigment derivatives of delphinidine and cyanidine[J]. Bull Soc Chim Biol.1967,49(4):395-413.
[24]Zaprometov M N, Bukhlaeva V I. Efficiency of use of various C14-precursors for the biosynthesis of flavonoids in the tea plant[J]. Biokhimiia.36(2):270.
[25]Zenk M H. Biosynthese von vanillin in Vanilla planifolia Andr[J]. Z. Pflanzenphysiol.1965,53: 404-414.
[26]Stafford H A. Possible multi-enzyme complexes regulating the formation of C6-C3 phenolic compounds and lignins in higher plants[J]. Rec. Adv. Phytochem.1974,8:53-79.
[27]Stafford H. Flavonoid evolution:an enzymic approach[J]. Plant Physiology.1991,96(3):680.
[28]方从兵,宛晓春,江昌俊.黄酮类化合物生物合成的研究进展(综述)[J].安徽农业大学学报,2005,32(4)：498-504.
[29]Christendat D, Saridakis V C, Turnbull J L. Use of Site-Directed Mutagenesis To Identify Residues Specific for Each Reaction Catalyzed by Chorismate Mutase-Prephenate Dehydrogenase from Escherichia coli[J]. Biochemistry.1998,37(45):15703-15712.
[30]Koch G L E, Shaw D C, Gibson F. Tyrosine biosynthesis in Aerobacter aerogenes:purification and properties of chorismate mutase-prephenate dehydrogenase[J]. Biochimica et Biophysica Acta (BBA)-Enzymology.1970,212(3):375-386.
[31]Stenmark S L, Pierson D L, Jensen R A, et al. Blue-green bacteria synthesise L-tyrosine by the pretyrosine pathway[J]. Nature (Lond).1974,247:290-292.
[32]Kim K H, Petersen M. Cloning and functional expression of hydroxyphenylpyruvate dioxygenase from Coleus blumei[J]. Phytochemistry.1997,45:1165-1172.
[33]petersen M, Simmonds M S J. Molecules of interest:rosmarinic acid[J]. Phytochemistry.2003, 62(2):121-125.
[34]孙视,刘晚苟.生态条件对银杏叶黄酮积累的影响[J].植物资源与环境.1998,7(3)：1-7.
[35]王义强.银杏枝叶生长规律及两次采叶试验研究[J].经济林研究.1995,13(4)：20-22.
[36]Lo S, Nicholson R. Reduction of light-induced anthocyanin accumulation in inoculated sorghum mesocotyls. Implications for a compensatory role in the defense response[J]. Plant Physiology. 1998,116(3):979.
[37]陈学森,张艳敏.叶用银杏资源评价及选优的研究[J].园艺学报.1997,24(3)：215-219.
[38]王英强,梁红.广东产银杏叶总黄酮含量变化[J].中药材.2001,24(4)：247-248.
[39]邢世岩,吴德军.叶用银杏种源,性别及无性系的因子和聚类分析[J].中南林学院学报.2000,20(2)：26-31.
[40]汪贵斌,曹福亮,方升佐等.银杏叶用园建园材料选择的研究[J].林业科学.2000,36(4)：26-31.
[41]刘德军,佘远国,汪鹏等.叶用银杏品种产量性状选择研究[J].经济林研究.2002,20(1)：30-31.
[42]王凌晖,曹福亮,汪贵斌等.银杏优质丰产园建园技术的研究进展[J].西北林学院学报.2005.20(1)：102-106.
[43]陈学森,章文才,邓秀新.树龄及季节对银杏叶黄酮与萜内酯含量的影响fJ].果树科学.1997,14(4)：226-229.
[44]程水源.影响银杏叶黄酮形成的主要因子及调控技术的研究[D].泰安：山东农业大学.2001
[45]Brinkman J, Boerner R. Nitrogen fertilization effects on foliar nutrient dynamics and autumnal resorption in maidenhair tree(Gingko biloba L.)[J]. Journal of Plant Nutrition.1994,17(2): 433-443.
[46]魏刚,赵洪亮.银杏不同营养器官中营养元素含量季节动态的研究[J].北京林业大学学报.1999,21(1)：96-99.
[47]王燕,程水源.提高银杏叶黄酮含量的调控措施[J].湖北农业科学.2002.40(5)：103-105.
[48]程水源,王燕,费永俊等.提高银杏叶黄酮含量的措施及其调控机理的研究[J].果树学报.2004,21(2)：116-119.
[49]冷平生,苏淑钗.光强与光质对银杏光合作用及黄酮苷与萜类内酯含量的影响[J].植物资源与环境学报.2002,11(1)：1-4.
[50]何丙辉,钟章成.不同光强与干旱胁迫对银杏枝叶构件生长的影响[J].广西师范大学学报：自然科学版.2005,23(3)：66-69.
[51]谢宝东,王华田.土壤水分含量对银杏叶黄酮和内酯含量的影响[J].山东林业科技.2002,(4)：1-3.
[52]景茂,曹宝亮,汪贵斌等.土壤水分含量对银杏生长及生物量分配的影响[J].南京林业大学学报：自然科学版.2005,29(3)：5-8.
[53]刘卫红,程水源.光照及机械损伤对银杏叶苯丙氨酸解氨酶活性的影响[J].湖北农业科学.2003,(3)：73-75.
[54]王燕,刘卫红,杜何为等.底物,末端产物对离体银杏叶苯丙氨酸解氨酶活性的影响[J].果树学报.2004,21(5)：443-446.
[55]王燕,李琳玲,许锋等.金属离子对盆栽银杏叶PAL酶活及黄酮含量的影响[J].南京林业大学学报：自然科学版.2007,31(2)：68-72.
[56]Xu F, Cheng S, Cheng S, et al. Time course of expression of chalcone synthase gene in Ginkgo biloba[J]. Plant Physiol. Mol. Biol.2007,33(4):309-317.
[57]Xu F, Cai R, Cheng S, et al. Molecular cloning, characterization and expression of phenylalanine ammonia-lyase gene from Ginkgo biloba[J]. African Journal of Biotechnology.2008,7(6): 721-729.
[58]Pang Y, Shen G, Wu W, et al. Characterization and expression of chalcone synthase gene from Ginkgo biloba[J]. Plant Science.2005,168(6):1525-1531.
[59]程水源,陈昆松,刘卫红等.植物苯丙氨酸解氨酶基因的表达调控与研究展望[J].果树学报.2003,20(5)：351-357.
[60]刘卫红.银杏叶苯丙氨酸解氨酶特性及其对叶黄酮含量调控的研究[D].武汉：华中农业大学, 2003
[61]Muir S R, Collins, G. J.,Robinson, S.,Hughes, S.,Bovy, A., Ric De Vos C H, van Tunen A J, et al. Overexpression of petunia chalcone isomerase in tomato results in fruit containing increased levels of flavonols[J]. Nat Biotechnol.2001,19(5):470-474.
[62]Li F X, Jin Z P, Zhao D X, et al. Overexpression of the Saussurea medusa chalcone isomerase gene in S. involucrata hairy root cultures enhances their biosynthesis of apigenin[J]. Phytochemistry. 2006,67(6):553-560.
[63]Harborne J, Williams C. Anthocyanins and other flavonoids[J]. Natural product reports.2001, 18(3):310-333.
[64]Ju Z, Liu C, Yuan Y. Activities of chalcone synthase and UDPGal:flavonoid-3-o-glycosy-ltransferase in relation to anthocyanin synthesis in apple[J]. Scientia Horticulturae.1995,63(3-4): 175-185.
[65]Welford R, Clifton I, Turnbull J, et al. Structural and mechanistic studies on anthocyanidin synthase catalysed oxidation of flavanone substrates:the effect of C-2 stereochemistry on product selectivity and mechanism[J]. Organic & Biomolecular Chemistry.2005,3(17):3117-3126.
[66]许锋.银杏GbPAL和GbANS基因的克隆与表达及ALA对类黄酮含量的影响[D].泰安：山东农业大学,2008
[67]Cosio E, McClure J. Kaempferol glycosides and enzymes of flavonol biosynthesis in leaves of a soybean strain with low photosynthetic rates[J]. Plant Physiology.1984,74(4):877.
[68]Saure M C. External control of anthocyanin formation in apple[J]. Scientia Horticulturae.1990, 42(3):181-218.
[69]El-Kereamy A, Chervin C, Roustan J, et al. Exogenous ethylene stimulates the long-term expression of genes related to anthocyanin biosynthesis in grape berries[J]. Physiologia Plantarum. 2003,119(2):175-182.
[70]Blankenship S, Unrath R. PAL and ethylene content during maturation of red and golden delicious apples[J]. Phytochemistry.1988,27(4):1001-1002.
[71]Awad M, de Jager A. Formation of flavonoids, especially anthocyanin and chlorogenic acid in [] Jonagold'apple skin:influences of growth regulators and fruit maturity [J]. Scientia Horticulturae. 2002,93(3-4):257-266.
[72]Hiratsuka S, Onodera H, Kawai Y, et al. ABA and sugar effects on anthocyanin formation in grape berry cultured in vitro [J]. Scientia Horticulturae.2001,90(1-2):121-130.
[73]刘叔倩,郑俊华.展望分子生物学技术银否研究中的应用[J].国外医药：植物约分册.1999,14(1)：4-6.
[74]Tulecke W. A Haploid Tissue Culture from the Female Gametophyte of Ginkgo biloba L[J]. Nature. 1964,203:94-95.
[75]Laurain D, Chenieux J, Tremouillaux-Guiller J. Direct embryogenesis from female haploid protoplasts of Ginkgo biloba L., a medicinal woody species[J]. Plant Cell Reports.1993,12(11): 656-660.
[76]张广辉,陈春秋.银杏离体培养生产次生代谢物研究进展[J].北京林业大学学报.2002,24(4)：130-134.
[77]陈学森,邓秀新,章文才.银杏组织培养与黄酮生产的研究[J].中国农业科学.1997,30(6)：55-60.
[78]陈学森,邓秀新.培养基及培养条件对银杏愈伤组织黄酮产量的影响[J].园艺学报.1997,24(4)：373-377.
[79]房建军,阙国宁.银杏愈伤组织生长和黄酮类化合物积累的关系[J].林业科学研究.1998,11(2)：124-129.
[80]孙天恩,李根保.银杏发根克隆及其培养技术研究进展[J].中草药.1998,29(增)：25-28.
[81]Winkel-Shirley B. Flavonoid biosynthesis. A colorful model for genetics, biochemistry, cell biology, and biotechnology[J]. Plant Physiology.2001,126(2):485-493.
[82]陈新,万德光,严铸云等.银杏苯丙氨酸解氨酶(PAL)的基因克隆[J].成都中医药大学学报.2004,27(1)：32-34.
[83]Pang Y, Shen G A, Liu C, et al. Molecular Cloning and Sequence Analysis of a Novel Chalcone Synthase cDNA from Ginkgo biloba[J]. Mitochondrial DNA.2004,15(4):283-290.
[84]Shen G, Pang Y, Wu W, et al. Cloning and characterization of a flavanone 3-hydroxylase gene from Ginkgo biloba[J]. Bioscience reports.2006,26(1):19-29.
[85]Shen G, Pang Y, Wu W, et al. Isolation and characterization of a putative anthocyanidin reductase gene from Ginkgo biloba[J]. Journal of plant physiology.2006,163(2):224-227.
[86]庞永珍.银杏黄酮和萜类化合物生物合成途径中重要相关基因的克隆和研究[D].上海：复旦大学,2005
[87]Bovy A, De Vos R, Kemper M, et al. High-flavonol tomatoes resulting from the heterologous expression of the maize transcription factor genes LC and C1[J]. The Plant Cell Online.2002, 14(10):2509.
[88]Verhoeyen M E, Bovy A, Collins G, et al. Increasing antioxidant levels in tomatoes through modification of the flavonoid biosynthetic pathway[J]. J. Exp. Bot.2002,53(377):2099-2106.
[89]Colliver S, Bovy A, Collins G, et al. Improving the nutritional content of tomatoes through reprogramming their flavonoid biosynthetic pathway[J]. Phytochemistry Reviews.2002,1(1): 113-123.
[90]Aharoni A, De Vos C, Wein M, et al. The strawberry FaMYB1 transcription factor suppresses anthocyanin and flavonol accumulation in transgenic tobacco[J]. The Plant Journal.2001,28(3): 319-332.
[91]路静,赵华燕,何奕昆等.高等植物启动子及其应用研究进展[J].自然科学进展.2004,14(8)：856-862.
[92]朱玉贤,李毅.现代分子生物学[M].北京：高等教育出版社,1997：17—59.
[93]夏江东,程在全,吴渝生等.高等植物启动子功能和结构研究进展[J].云南农业大学学报.2006,21(1)：7-14.
[94]张春晓,王文棋,蒋湘宁等.植物基因启动子研究进展[J].遗传学报.2004,31(12)：1455-1464.
[95]Zhang C X, Wang W Q, Jiang X N, et al. Review on plant gene promoters[J]. Acta genetica Sinica. 2004,31(12):1455-1464.
[96]Carey M F, Smale S T. Transcriptional regulation in eukaryotes:concepts, strategies, and techniques[M]. Cold Spring Harbor Laboratory Pr,2000:
[97]Gatz C, Lenk I. Promoters that respond to chemical inducers[J]. Trends in plant science.1998,3(9): 352-358.
[98]Zuo J, Chua N H. Chemical-inducible systems for regulated expression of plant genes[J]. Current Opinion in Biotechnology.2000,11(2):146-151.
[99]Weinmann P, Gossen M, Hillen W, et al. A chimeric transactivator allows tetracycline-responsive gene expression in whole plants[J]. The Plant Journal.2003,5(4):559-569.
[100]Zhu Q, Dabi T, Lamb C. TATA box and initiator functions in the accurate transcription of a plant minimal promoter in vitro[J]. The Plant Cell Online.1995,7(10):1681-1689.
[101]Neve R L, West R W, Rodriguez R L. Eukaryotic DNA fragments which act as promoters for a plasmid gene[J]. Natur.1979,277(5694):324-325.
[102]Sidhu R S, Mathewes S, Bollon A P. Selection of secretory protein-encoding genes by fusion with PHO5 in Saccharomyces cerevisiae[J]. Gene.1991,107(1):111-118.
[103]李姗姗,迟彦,李凌飞等.启动子克隆方法研究进展[J].中国生物工程杂志.2005,25(7)：9-16.
[104]Vida T A, Graham T R, Emr S D. In vitro reconstitution of intercompartmental protein transport to the yeast vacuole[J]. Journal of Cell Biology.1990,111(6):2871-2884.
[105]Friedrich G, Soriano P. Promoter traps in embryonic stem cells:a genetic screen to identify and mutate developmental genes in mice[J]. Genes Dev.1991,5(9):1513-1523.
[106]Lindsey K, Wei W, Clarke M C, et al. Tagging genomic sequences that direct transgene expression by activation of a promoter trap in plants[J]. Transgenic research.1993,2(1):33-47.
[107]王爱民,陈石,燕一等.Ac/Ds (GUS)介导的水稻启动子捕获系统的建立[J].植物生理与分子生物学学报.2005,31(6)：575-580.
[108]方华舟,涂知明.水稻花粉组织特异性启动子捕获实验[J].生物学杂志.2007,24(5)：37-39.
[1091李竹红,刘德培,梁植权.改进的反向PCR技术克隆转移基因的旁侧序列[J].生物化学与生物物理进展.1999,26(6)：600-602.
[110]Triglia T, Peterson M, Kemp D. A procedure for in vitro amplification of DNA segments that lie outside the boundaries of known sequences[J]. Nucleic Acids Research.1988,16(16):8186.
[111]Jones D H, Winistorfer S C. Recombinant circle PCR and recombination PCR for site-specific mutagenesis without PCR product purification[J]. BioTechniques.1992,12(4):528-535.
[112]Jones D H, Winistorfer S C. Amplification of 4-9-kb human genomic DNA flanking a known site using a panhandle PCR variant[J]. BioTechniques.1997,23(1):132-138.
[113]Prashar Y, Weissman S M. Analysis of differential gene expression by display of 3'end restriction fragments of cDNAs[J]. Proceedings of the National Academy of Sciences.1996,93(2):659-663.
[114]Shyamala V, Ames G F. Amplification of bacterial genomic DNA by the polymerase chain reaction and direct sequencing after asymmetric amplification:application to the study of periplasmic permeases[J]. Journal of bacteriology.1989,171(3):1602-1608.
[115]Han J, Luhs W, Sonntag K, et al. Functional characterisation of p-ketoacyl-CoA synthase genes from Brassica napus [J]. L. Plant Mol. Biol.2001,46:229-239.
[116]王新国,张国华.用衔接头PCR克隆新的胡萝卜Ⅱ型转化酶基因启动子[J].中国生物化学与分子生物学报.2001,17(1)：61-65.
[117]Liu Y G, Whittier R F. Thermal asymmetric interlaced PCR:automatable amplification and sequencing of insert end fragments from P1 and YAC clones for chromosome walking [J]. Genomics.1995,25(3):674-681.
[118]Chen S, Jin W, Wang M, et al. Distribution and characterization of over 1000 T-DNA tags in rice genome[J]. The Plant Journal.2003,36(1):105-113.
[119]Zhang Q. Strategies for developing green super rice[J]. Proceedings of the National Academy of Sciences.2007,104(42):16402-16409.
[120]Schmid C D, Praz V, Delorenzi M, et al. The Eukaryotic Promoter Database EPD:the impact of in silico primer extension[J]. Nucleic Acids Research.2004,32:82-85.
[121]Heinemeyer T, Wingender E, Reuter I, et al. Databases on transcriptional regulation:TRANSFAC, TRRD and COMPEL[J]. Nucleic Acids Research.1998,26(1):362-367.
[122]Higo K, Ugawa Y, Iwamoto M, et al. Plant cis-acting regulatory DNA elements (PLACE) database:1999[J]. Nucleic Acids Research.1999,27(1):297-300.
[123]Rombauts S, Dehais P, Van Montagu M, et al. PlantCARE, a plant cis-acting regulatory element database[J]. Nucleic Acids Research.1999,27(1):295-296.
[124]朱廷恒,罗红丽,宋凤鸣等.水稻病程相关蛋白基因OsPR-4b启动子的克隆及缺失体构建fJl.浙江大学学报：农业与生命科学版,2005,31(1)：22-26.
[125]杨英军,周鹏.番木瓜proteinase omega基因启动子的克隆及功能初步研究[J].云南植物研究.2005,27(5)：545-551.
[126]宗成文,章镇,房经贵等.葡萄LEAFY基因启动子的克隆与序列分析[J].南京农业大学学报.2007,30(4)：20-25.
[127]Jefferson R A, Kavanagh T A, Bevan M W. GUS fusions:beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants[J]. The EMBO Journal.1987,6(13):3901-3907.
[128]Baurle I, Laux T. Regulation of WUSCHEL transcription in the stem cell niche of the Arabidopsis shoot meristem[J]. The Plant Cell Online.2005,17(8):2271-2280.
[129]Matarasso N, Schuster S, Avni A. A novel plant cysteine protease has a dual function as a regulator of 1-aminocyclopropane-l-carboxylic acid synthase gene expression[J]. The Plant Cell Online.2005,17(4):1205-1216.
[130]王新力,彭学贤.香蕉果实成熟相关基因ACO1启动子区的克隆及其功能初探[J].生物工程学报.2001,17(4)：428-431.
[131]Atkinson R G, Bolitho K M, Wright M A, et al. Apple ACC-oxidase and polygalacturonase: ripening-specific gene expression and promoter analysis in transgenic tomato[J]. Plant Molecular Biology.1998,38(3):449-460.
[132]Wang Z Y, MacRae E A, Wright M A, et al. Polygalacturonase gene expression in kiwifruit: relationship to fruit softening and ethylene production[J]. Plant Molecular Biology.2000,42(2): 317-328.
[133]袁正强,吴家和.三个韧皮部特异性启动子在转基因烟草中表达的比较研究[J].农业生物技术学报.2002,10(1)：6-9.
[134]Melegari M, Scaglioni P P, Wands J R. Cloning and characterization of a novel hepatitis B virus x binding protein that inhibits viral replication[J]. Journal of virology.1998,72(3):1737-1743.
[135]Liao Y, Zou H F, Wei W, et al. Soybean GmbZIP44, GmbZIP62 and GmbZIP78 genes function as negative regulator of ABA signaling and confer salt and freezing tolerance in transgenic Arabidopsis[J]. Planta.2008,228(2):225-240.
[136]Wang S. Identification of novel pathogen-responsive cis-elements and their binding proteins in the promoter of OsWRKY13, a gene regulating rice disease resistance[J]. Plant, Cell and Environment.2007,31:86-96.
[137]Galas D J, Schmitz A. DNase I footprinting; a simple method for the detection of protein-DNA binding specificty[J]. Nucleic Acids Res.1978,5:3157-3170.
[138]Ackers G, Johnson A, Shea M. Quantitative model for gene regulation by lambda phage repressor[J]. Proc Natl Acad Sci USA.1982,79(4):1129-1136.
[139]Nakamura I, Koike K. Identification of a binding protein to the X gene promoter region of hepatitis B virus[J]. Virology.1992,191(2):533-540.
[140]Bouffartigues E, Buckle M, Badaut C, et al. H-NS cooperative binding to high-affinity sites in a regulatory element results in transcriptional silencing[J]. Nat Struct Mol Biol.2007,14(5): 441-448.
[141]Adams B, Cha H, Cleary J, et al. DEK binding to class Ⅱ MHC Y-box sequences is gene-and allele-specific[J]. Arthritis Res Ther. 2003,5(4):226-233.
[142]Flanigan A, Gardner J F. Interaction of the Gifsy-1 Xis protein with the Gifsy-1 attP sequence[J]. Journal of bacteriology.2007,189(17):6303-6311.
[143]Smith J V, Luo Y. Studies on molecular mechanisms of Ginkgo biloba extract[J]. Applied microbiology and biotechnology.2004,64(4):465-472.
[144]Mehdy M, Lamb C. Chalcone isomerase cDNA cloning and mRNA induction by fungal elicitor, wounding and infection[J]. The EMBO Journal.1987,6(6):1527.
[145]van Tunen A J, Koes R E, Spelt C E, et al. Cloning of the two chalcone flavanone isomerase genes from Petunia hybrida:coordinate, light-regulated and differential expression of flavonoid genes[J]. EMBO J.1988,7(5):1257-1263.
[146]Blyden E R, Doerner P W, Lamb C J, et al. Sequence analysis of a chalcone isomerase cDNA of Phaseolus vulgaris L[J]. Plant Mol Biol.1991,16(1):167-169.
[147]Grotewold E, Peterson T. Isolation and characterization of a maize gene encoding chalcone flavonone isomerase[J]. Mol Gen Genet.1994,242(1):1-8.
[148]Wood A J, Davies E. A cDNA Encoding Chalcone Isomerase from Aged Pea Epicotyls[J]. Plant Physiology.1994,104(4):1465-1466.
[149]McKhann H I, Hirsch A M. Isolation of chalcone synthase and chalcone isomerase cDNAs from alfalfa (Medicago sativa L.):highest transcript levels occur in young roots and root tips[J]. Plant Mol Biol.1994,24(5):767-777.
[150]Li F, Jin Z, Qu W, et al. Cloning of a cDNA encoding the Saussurea medusa chalcone isomerase and its expression in transgenic tobacco[J]. Plant Physiol Biochem.2006,44(7-9):455-461.
[151]Gensheimer M, Mushegian A. Chalcone isomerase family and fold:No longer unique to plants [J]. Protein Sci.2004,13(2):540-544.
[152]Druka A, Kudrna D, Rostoks N, et al. Chalcone isomerase gene from rice (Oryza sativa) and barley (Hordeum vulgare):physical, genetic and mutation mapping[J]. Gene.2003,302(1-2): 171-178.
[153]Shimada N, Aoki T, Sato S, et al. A Cluster of genes encodes the two types of chalcone isomerase involved in the biosynthesis of general flavonoids and legume-specific 5-deoxy(iso)flavonoids in Lotus japonicus[J]. Plant Physiology.2003,131(3):941-951.
[154]McClean P E, Lee R K. Genetic architecture of chalcone isomerase non-coding regions in common bean (Phaseolus vulgaris L.)[J]. Genome.2007,50(2):203-214.
[155]Kuittinen H, Aguade M. Nucleotide variation at the chalcone isomerase locus in Arabidopsis thaliana[J]. Genetics.2000,155(2):863-872.
[156]Van Tunen A, Mur L, Brouns G, et al. Pollen-and anther-specific chi promoters from petunia: tandem promoter regulation of the chiA gene[J]. The Plant Cell Online.1990,2(5):393-401.
[157]程水源,陈昆松,杜何为等.银杏RNA的提取[J].果树学报.2005,22(4)：428-429.
[158]Sambrook J, Russell D W. Molecular cloning:a laboratory manual[M]. CSHL press,2001:
[159]魏春红,李毅.现代分子生物学实验[M].北京：高等教育出版社,2006：135-137.
[160]Jansson S, Meyer-Gauen G, Cerff R, et al. Nucleotide distribution in gymnosperm nuclear sequences suggests a model for GC-content change in land-plant nuclear genomes[J]. Journal of molecular evolution.1994,39(1):34-46.
[161]Jez J M, Noel J P. Reaction mechanism of chalcone isomerase:pH dependence, diffusion control, and product binding differences [J]. J. Biol. Chem.2002,277(2):1361-1369.
[162]Fouche S D, Dubery I A. Chalcone isomerase from Citrus sinensis:purification and characterization[J]. Phytochemistry.1994,37(1):127-132.
[163]Bradford M M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding[J]. Analytical biochemistry.1976,72(1-2): 248-254.
[164]Moustafa E, Wong E. Purification and properties of chalcone-flavanone isomerase from soya bean seed[J]. Phytochemistry.1967,6(5):625-632.
[165]程水源,王燕.银杏叶黄酮含量变化及分布规律的研究[J].园艺学报.2001,28(4)：353-355.
[166]Lutcke H A, Chow K C, Mickel F S, et al. Selection of AUG initiation codons differs in plants and animals[J]. EMBO J.1987,6(1):43-48.
[167]Folter S, Angenent G. trans meets cis in MADS science[J]. Trends in plant science.2006,11(5): 224-231.
[168]Yang T, Poovaiah B. A calmodulin-binding/CGCG box DNA-binding protein family involved in multiple signaling pathways in plants[J]. Journal of Biological Chemistry.2002,277 (47): 45049-45058.
[169]Piechulla B, Merforth N, Rudolph B. Identification of tomato Lhc promoter regions necessary for circadian expression[J]. Plant Molecular Biology.1998,38(4):655-662.
[170]Chen C, Chen Z. Potentiation of developmentally regulated plant defense response by AtWRKY18, a pathogen-induced Arabidopsis transcription factor[J]. Plant Physiology.2002, 129(2):706-716.
[171]Schunmann P, Richardson A, Vickers C, et al. Promoter analysis of the barley Phtl; 1 phosphate transporter gene identifies regions controlling root expression and responsiveness to phosphate deprivation[J]. Plant Physiology.2004,136(4):4205-4214.
[172]Dunn M, White A, Vural S, et al. Identification of promoter elements in a low-temperature-responsive gene (blt4.9) from barley(Hordeum vulgare L.)[J]. Plant Molecular Biology.1998, 38(4):551-564.
[173]Hartmann U, Sagasser M, Mehrtens F, et al. Differential combinatorial interactions of cis-acting elements recognized by R2R3-MYB, BZIP, and BHLH factors control light-responsive and tissue-specific activation of phenylpropanoid biosynthesis genes[J]. Plant Mol Biol.2005,57(2): 155-171.
[174]Bate N, Twell D. Functional architecture of a late pollen promoter:pollen-specific transcription is developmentally regulated by multiple stage-specific and co-dependent activator elements[J]. Plant Molecular Biology.1998,37(5):859-869.
[175]Yoshioka S, Taniguchi F, Miura K, et al. The novel Myb transcription factor LCR1 regulates the CO2-responsive gene Cahl, encoding a periplasmic carbonic anhydrase in Chlamydomonas reinhardtii[J]. Plant Cell.2004,16(6):1466-1477.
[176]Solano R, Nieto C, Avila J, et al. Dual DNA binding specificity of a petal epidermis-specific MYB transcription factor (MYB. Ph3) from Petunia hybrida[J]. The EMBO Journal.1995,14(8): 1773-1784.
[177]Lam E, Chua N. ASF-2:a factor that binds to the cauliflower mosaic virus 35S promoter and a conserved GATA motif in Cab promoters[J]. The Plant Cell Online.1989,1(12):1147-1156.
[178]Lopez-Molina L, Chua N H. A null mutation in a bZIP factor confers ABA-insensitivity in Arabidopsis thaliana[J]. Plant and Cell Physiology.2000,41(5):541-547.
[179]Ogawa M, Hanada A, Yamauchi Y, et al. Gibberellin biosynthesis and response during Arabidopsis seed germination[J]. The Plant Cell Online.2003,15(7):1591-1604.
[180]Boyle B, Brisson N. Repression of the defense gene PR-10a by the single-stranded DNA binding protein SEBF[J]. The Plant Cell Online.2001,13(11):2525-2537.
[181]Yamagata H, Yonesu K, Hirata A, et al. TGTCACA Motif Is a Novel cis-Regulatory Enhancer Element Involved in Fruit-specific Expression of thecucumisin Gene[J]. Journal of Biological Chemistry.2002,277(13):11582-11590.
[182]Zhou D. Regulatory mechanism of plant gene transcription by GT-elements and GT-factors[J]. Trends in plant science.1999,4(6):210-214.
[183]Park H, Kim M, Kang Y, et al. Pathogen-and NaCl-induced expression of the SCaM-4 promoter is mediated in part by a GT-1 box that interacts with a GT-1-like transcription factor[J]. Plant Physiology.2004,135(4):2150-2161.
[184]Rawat R, Xu Z-F, Yao K-M, et al. Identification of cis-elements for ethylene and circadian regulation of the Solanum melongena gene encoding cysteine proteinase[J]. Plant Mol Biol.2005, 57(5):629-643.
[185]Itzhaki H, Maxson J M, Woodson W R. An ethylene-responsive enhancer element is involved in the senescence-related expression of the carnation glutathione-S-transferase (GST1) gene[J]. PNAS.1994,91(19):8925-8929.
[186]Montgomery J, Goldman S, Deikman J, et al. Identification of an ethylene-responsive region in the promoter of a fruit ripening gene[J]. PNAS.1993,90(13):5939-5943.
[187]Jez J M, Bowman M E, Dixon R A, et al. Structure and mechanism of the evolutionarily unique plant enzyme chalcone isomerase[J]. Nat Struct Biol.2000,7(9):786-791.
[188]Bednar R, Hadcock J. Purification and characterization of chalcone isomerase from soybeans[J]. Journal of Biological Chemistry.1988,263(20):9582-9588.
[189]Dixon R, Richard Blyden E, Robbins M, et al. Comparative biochemistry of chalcone isomerases[J]. Phytochemistry.1988,27(9):2801-2808.
[190]Shirley B, Kubasek W, Storz G, et al. Analysis of Arabidopsis mutants deficient in flavonoid biosynthesis[J]. The Plant Journal.1995,8(5):659-671.
[191]Xu F, Cheng S Y, Cheng S H, et al. Time course of expression of chalcone synthase gene in Ginkgo biloba[J]. Journal of Plant Physiology and Molecular Biology.2007,33(4):309-317.
[192]Li J, Ou-Lee T, Raba R, et al. Arabidopsis flavonoid mutants are hypersensitive to UV-B irradiation[J]. The Plant Cell Online.1993,5(2):171-179.
[193]Bieza K, Lois R. An Arabidopsis mutant tolerant to lethal ultraviolet-B levels shows constitutively elevated accumulation of flavonoids and other phenolics[J]. Plant Physiology.2001,126(3): 1105-1115.
[194]Kush A, Goyvaerts E, Chye M L, et al. Laticifer-specific gene expression in Hevea brasiliensis (rubber tree)[J]. Proceedings of the National Academy of Sciences.1990,87(5):1787-1790.
[195]Ardi R, Kobiler I, Jacoby B, et al. Involvement of epicatechin biosynthesis in the activation of the mechanism of resistance of avocado fruits to Colletotrichum gloeosporioides[J]. Physiological and Molecular Plant Pathology.1998,53(5-6):269-285.
[196]Abe H, Yamaguchi-Shinozaki K, Urao T, et al. Role of Arabidopsis MYC and MYB homologs in drought-and abscisic acid-regulated gene expression[J]. The Plant Cell Online.1997,9(10): 1859-1868.
[197]蔡葛平,郭燕红,姚辉等.矮壮素和赤霉素对黄芩生物量及根中黄酮类成分产量的影响[J].中国农学通报.2008,24(7)：213-217.
[198]程水源,王燕,李俊凯等.内源激素含量与银杏叶中类黄酮含量的关系[J].林业科学.2004,40(6)：45-49.
[199]Von Wettstein D, Gough S, Kannangara C. Chlorophyll biosynthesis[J]. The Plant Cell.1995,7(7): 1039-1057.
[200]汪良驹,王中华,李志强等.5-氨基乙酰丙酸促进苹果果实着色的效应[J].果树学报.2004,21(006)：512-515.
[201]程水源,王燕,李俊凯等.银杏叶片色素含量与黄酮含量关系的研究[J].林业科学.2001,37(5)：31-34.
[202]Brugliera F, Barri-Rewell G, Holton T, et al. Isolation and characterization of a flavonoid 3'-hydroxylase cDNA clone corresponding to the Htl locus of Petunia hybrida[J]. The Plant Journal.2002,19(4):441-451.
[203]Seitz C, Eder C, Deiml B, et al. Cloning, functional identification and sequence analysis of flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase cDNAs reveals independent evolution of flavonoid 3',5'-hydroxylase in the Asteraceae family[J]. Plant Molecular Biology.2006,61(3): 365-381.
[204]Tanaka Y. Flower colour and cytochromes P450[J]. Phytochemistry Reviews.2006,5(2): 283-291.
[205]Nakatsuka T, Nishihara M, Mishiba K, et al. Heterologous expression of two gentian cytochrome P450 genes can modulate the intensity of flower pigmentation in transgenic tobacco plants[J]. Molecular Breeding.2006,17(2):91-99.
[206]Jeong S, Goto-Yamamoto N, Hashizume K, et al. Expression of the flavonoid 3'-hydroxylase and flavonoid 3',5'-hydroxylase genes and flavonoid composition in grape (Vitis vinifera)[J]. Plant
Science.2006,170(1):61-69.
[207]Kaplan B, Davydov O, Knight H, et al. Rapid transcriptome changes induced by cytosolic Ca2+ transients reveal ABRE-related sequences as Ca2+-responsive cis elements in Arabidopsis[J]. Plant Cell.2006,18(10):2733-2748.
[208]Luo H, Song F, Goodman R M, et al. Up-regulation of OsBIHD1, a rice gene encoding BELL homeodomain transcriptional factor, in disease resistance responses[J]. Plant Biology.2005,7(5): 459-468.
[209]Abe H, Urao T, Ito T, et al. Arabidopsis AtMYC2 (bHLH) and AtMYB2 (MYB) function as transcriptional activators in abscisic acid signaling[J]. The Plant Cell Online.2003,15(1):63-67.
[210]Fujiwara T, Beachy R N. Tissue-specific and temporal regulation of a β-conglycinin gene:roles of the RY repeat and other cis-acting elements[J]. Plant Molecular Biology.1994,24(2):261-272.
[211]Plesch G, Ehrhardt T, Mueller-Roeber B. Involvement of TAAAG elements suggests a role for Dof transcription factors in guard cell-specific gene expression[J]. The Plant Journal.2002,28(4): 455-464.
[212]Kram B, Xu W, Carter C. Uncovering the Arabidopsis thaliana nectary transcriptome: investigation of differential gene expression in floral nectariferous tissues[J]. BMC plant biology. 2009,9(1):92.
[213]Tang W, Perry S. Binding site selection for the plant MADS domain protein AGL15[J]. Journal of Biological Chemistry.2003,278(30):28154.
[214]Ellerstr m M, St lberg K, Ezcurra I, et al. Functional dissection of a napin gene promoter: identification of promoter elements required for embryo and endosperm-specific transcription[J]. Plant Molecular Biology.1996,32(6):1019-1027.
[215]Fusada N, Masuda T, Kuroda H, et al. Identification of a novel cis-element exhibiting cytokinin-dependent protein binding in vitro in the 5'-region of NADPH-protochlorophyllide oxidoreductase gene in cucumber[J]. Plant Molecular Biology.2005,59(4):631-645.
[216]Kim S, Chung H, Thomas T. Isolation of a novel class of bZIP transcription factors that interact with ABA-responsive and embryo-specification elements in the Dc3 promoter using a modified yeast one-hybrid system[J]. The Plant Journal.2002,11(6):1237-1251.
[217]Eulgem T, Rushton P, Robatzek S, et al. The WRKY superfamily of plant transcription factors[J]. Trends in plant science.2000,5(5):199-205.
[218]Baumann K, De Paolis A, Costantino P, et al. The DNA binding site of the Dof protein NtBBF1 is essential for tissue-specific and auxin-regulated expression of the rolB oncogene in plants [J]. The Plant Cell Online.1999,11(3):323-334.
[219]Yamazaki S, Sato K, Suhara K, et al. Importance of the proline-rich region following signal-anchor sequence in the formation of correct conformation of microsomal cytochrome P450s[J]. Journal of biochemistry.1993,114(5):652-657.
[220]Murakami K, Mihara K, Omura T. The transmembrane region of microsomal cytochrome P450 identified as the endoplasmic reticulum retention signal[J]. Journal of biochemistry.1994,116(1): 164-173.
[221]Chapple C. Molecular-genetic analysis of plant cytochrome P450-dependent monooxygenases[J]. Annual Review of Plant Biology.1998,49(1):311-343.
[222]许志茹,崔国新,李春雷等.芜菁的类黄酮3'羟化酶基因克隆和UV-A诱导表达特性[J].植物生理学通讯.2008,44(5)：931-935.
[223]Xu B, Li J, Zhang X, et al. Cloning and molecular characterization of a functional flavonoid 3'-hydroxylase gene from Brassica napus[J]. Journal of plant physiology.2007,164(3):350-363.
[224]Schoenbohm C, Martens S, Eder C, et al. Identification of the Arabidopsis thaliana flavonoid 3'-hydroxylase gene and functional expression of the encoded P450 enzyme[J]. Biological Chemistry.2000,381(8):749-754.
[225]Xu F, Cheng H, Cai R, et al. Molecular cloning and function analysis of an anthocyanidin synthase gene from Ginkgo biloba, and its expression in abiotic stress responses[J]. Mol. Cell. 2008,26(6):536-547.
[226]Kreuzaler F, Ragg H, Fautz E, et al. UV-induction of chalcone synthase mRNA in cell suspension cultures of Petroselinum hortense[J]. Proceedings of the National Academy of Sciences of the United States of America.1983,80(9):2591.
[227]Koes R, Quattrocchio F, Mol J. The flavonoid biosynthetic pathway in plants:function and evolution[J]. BioEssays.2005,16(2):123-132.
[228]Deikman J, Hammer P E. Induction of anthocyanin accumulation by cytokinins in Arabidopsis thaliana[J]. Plant Physiology.1995,108(1):47-57.
[229]Herrmann K M, Weaver L M. The shikimate pathway[J]. Annual Review of Plant Physiology and Plant Molecular Biology.1999,50(1):473-503.
[230]Kumar A, Ellis B E. The phenylalanine ammonia-lyase gene family in raspberry. Structure, expression, and evolution[J]. Plant Physiology.2001,127(1):230-239.
[231]Hermann K M. The shikimate pathway:early steps in the biosynthesis of aromatic compounds[J]. Plant Cell.1995,7(3):907-919.
[232]Xujun W, Xiaoli W, Xugang L, et al. Isolation of rice EPSP synthase cDNA and its sequence analysis and copy number determination[J]. Acta Botanica Sinica.2002,44(2):188-192.
[233]Klee H J, Muskopf Y M, Gasser C S. Cloning of an Arabidopsis thaliana gene encoding 5-enolpyruvylshikimate-3-phosphate synthase:sequence analysis and manipulation to obtain glyphosate-tolerant plants[J]. Molecular and General Genetics.1987,210(3):437-442.
[234]刘东军,张锐,郭三堆等.棉花品系Y18在草甘膦胁迫下的epsps基因表达分析研究[J].中国生物工程杂志.2008,28(10)：55-59.
[235]蒋向,戴雄泽,李育强等.薤白EPSPs基因在不同组织表达的半定量分析[J].湖南农业大学学报(自然科学版).2007,33(5)：542-545.
[236]Gong Y, Liao Z, Chen M, et al. Characterization of 5-enolpyruvylshikimate 3-phosphate synthase gene from Camptotheca acuminata[J]. Biologia Plantarum.2006,50(4):542-550.
[237]Papanikou E, Brotherton J E, Widholm J M. Length of time in tissue culture can affect the selected glyphosate resistance mechanism[J]. Planta.2004,218(4):589-598.
[238]Stallings W C, Abdel-Meguid S S, Lim L W, et al. Structure and topological symmetry of the glyphosate target 5-enolpyruvylshikimate-3-phosphate synthase:a distinctive protein fold[J]. Proceedings of the National Academy of Sciences.1991,88(11):5046-5050.
[239]Shirley-Winkel B. It takes a garden, How work on diverse plant species ha contributed to an understanding of flavonoid metabolism[J]. Plant Physiology.2001,127(2316):1399-1404.
[240]van der Krol A R, van Poecke R M P, Vorst O F J, et al. Developmental and wound-, cold-, desiccation-, ultraviolet-B-stress-induced modulations in the expression of the petunia zinc finger transcription factor gene ZPT2-2[J]. Plant Physiology.1999,121(4):1153-1162.
[241]Benfey P N, Takatsuji H, Ren L, et al. Sequence requirements of the 5-enolpyruvylshikimate-3-phosphate synthase 5 [prime]-upstream region for tissue-specific expression in flowers and seedlings[J]. Plant Cell.1990,2(9):849-856.
[242]魏建华,宋艳茹.木质素生物合成途径及调控的研究进展[J].植物学报：英文版.2001,43(008)：771-779.
[243]Lacombe E, Hawkins S, Van Doorsselaere J, et al. Cinnamoyl CoA reductase, the first committed enzyme of the lignin branch biosynthetic pathway:cloning, expression and phylogenetic relationships[J]. The Plant Journal.2003,11(3):429-441.
[244]Piquemal J, Lapierre C, Myton K, et al. Down-regulation of cinnamoyl-CoA reductase induces significant changes of lignin profiles in transgenic tobacco plants[J]. The Plant Journal.2002, 13(1):71-83.
[245]Jones L, Ennos A R, Turner S R. Cloning and characterization of irregular xylem4 (irx4):a severely lignin-deficient mutant of Arabidopsis[J]. The Plant Journal.2001,26(2):205-216.
[246]李波,梁颖,柴友荣.植物肉桂酰辅酶A还原酶(CCR)基因的研究进展[J].分子植物育种.2006,4(3)：55-65.
[247]Sarni F, Grand C, Boudet A. Purification and properties of cinnamoyl-CoA reductase and cinnamyl alcohol dehydrogenase from poplar stems (Populus×euramericana)[J]. European Journal of Biochemistry.2005,139(2):259-265.
[248]Goffner D, Campbell M, Campargue C, et al. Purification and characterization of cinnamoyl-coenzyme A:NADP oxidoreductase in Eucalyptus gunnii[J]. Plant Physiology.1994, 106(2):625-632.
[249]Luderitz T, Grisebach H. Enzymic synthesis of lignin precursors comparison of cinnamoyl-CoA reductase and cinnamyl alcohol:NADP+ dehydrogenase from spruce (Picea abies L.) and soybean (Glycine max L.)[J]. European Journal of Biochemistry.2005, 119(1):115-124.
[250]McInnes R, Lidgett A, Lynch D, et al. Isolation and characterization of a cinnamoyl-CoA reductase gene from perennial ryegrass (Lolium perenne)[J]. Journal of plant physiology.2002, 159(4):415-422.
[251]Larsen K. Cloning and characterization of a ryegrass (Lolium perenne) gene encoding cinnamoyl-CoA reductase (CCR) [J]. Plant Science.2004,166(3):569-581.
[252]Lewis N, Davin L. Evolution of lignan and neolignan biochemical pathways:American chemical society [J]. Isopentenoids and other natural products:evolution and function.1994,202-246.
[253]Li L, Cheng X, Lu S, et al. Clarification of cinnamoyl co-enzyme A reductase catalysis in monolignol biosynthesis of aspen[J]. Plant and Cell Physiology.2005,46(7):1073-1082.
[254]Lauvergeat V, Lacomme C, Lacombe E, et al. Two cinnamoyl-CoA reductase (CCR) genes from Arabidopsis thaliana are differentially expressed during development and in response to infection with pathogenic bacteria [J]. Phytochemistry.2001,57(7):1187-1195.
[255]Pichon M, Courbou I, Beckert M, et al. Cloning and characterization of two maize cDNAs encoding cinnamoyl-CoA reductase (CCR) and differential expression of the corresponding genes[J]. Plant Molecular Biology.1998,38(4):671-676.
[256]Wengenmayer H, Ebel J, Grisebach H. Purification and properties of a cinnamoyl CoA:NADPH reductase from cell suspension cultures of soybean (Glycine max) [J]. European Journal of Biochemistry.65(2):529-536.
[257]Lacombe E, Van Doorsselaere J, Boerjan W, et al. Characterization of cis-elements required for vascular expression of the cinnamoyl CoA reductase gene and for protein-DNA complex formation[J]. The Plant Journal.2000,23(5):663-676.
[258]Selman-Housein G, Lopez M, Hernandez D, et al. Molecular cloning of cDNAs coding for three sugarcane enzymes involved in lignification[J]. Plant Sci.1999,143(2):163-171.
[259]Raven J. The evolution of vascular land plants in relation to supracellular transport processes[J]. Advances in botanical research.1977,5(1):153-219.
[260]van Beek T. Chemical analysis of Ginkgo biloba leaves and extracts[J]. Journal of Chromatography A.2002,967(1):21-55.
[261]Heller W, Hahlbrock K. Highly purified" flavanone synthase" from parsley catalyzes the formation of naringenin chalcone[J]. Archives of biochemistry and biophysics.1980,200(2): 617-619.
[262]许锋.银杏查尔酮合成酶基因和苯丙氨酸解氨酶基因的克隆及表达[D].武汉：华中农业大学,2005
[263]Dynan W. Modularity in promoters and enhancers[J]. Cell.1989,58(1):1-4.
[264]Loake G J, Faktor O, Lamb C J, et al. Combination of H-box [CCTACC (N) 7CT] and G-box (CACGTG) cis elements is necessary for feed-forward stimulation of a chalcone synthase promoter by the phenylpropanoid-pathway intermediate p-coumaric acid[J]. PNAS.1992,89(19): 9230-9234.
[265]Schulze-Lefert P, Becker-Andre M, Schulz W, et al. Functional architecture of the light-responsive chalcone synthase promoter from parsley[J]. Plant Cell.1989,1(7):707-714.
[266]Hartmann U, Valentine W J, Christie J M, et al. Identification of UV/blue light-response elements in the Arabidopsis thaliana chalcone synthase promoter using a homologous protoplast transient expression system[J]. Plant Molecular Biology.1998,36(5):741-754.
[267]Lawton M A, Dean S M, Dron M, et al. Silencer region of a chalcone synthase promoter contains multiple binding sites for a factor, SBF-1, closely related to GT-1[J]. Plant Molecular Biology. 1991,16(2):235-249.
[268]van der Meer I M, Brouwer M, Spelt C E, et al. The TACPyAT repeats in the chalcone synthase promoter of Petunia hybrida act as a dominant negative cis-acting module in the control of organ-specific expression[J]. The Plant Journal.1992,2(4):525-535.
[269]Wang J, Letham D S, Cornish E, et al. Studies of cytokinin action and metabolism using tobacco plants expressing either the ipt or the GUS gene controlled by a chalcone synthase Promoter. I. Developmental Features of the Transgenic Plants[J]. Functional Plant Biology.1997,24(5): 661-672.
[270]Faktor O, Kooter J M, Dixon R A, et al. Functional dissection of a bean chalcone synthase gene promoter in transgenic tobacco plants reveals sequence motifs essential for floral expression[J]. Plant Molecular Biology.1996,32(5):849-859.
[271]许锋,程水源,王燕等.TAIL-PCR方法快速克隆银杏查尔酮合成酶基因及序列分析[J].果树学报.2007,24(2)：237-243.
[272]Jefferson R A. Assaying chimeric genes in plants:the GUS gene fusion system[J]. Plant Molecular Biology Reporter.1987,5(4):387-405.
[273]Thum K E, Kim M, Morishige D T, et al. Analysis of barley chloroplast psbD light-responsive promoter elements in transplastomic tobacco[J]. Plant Molecular Biology.2001,47(3):353-366.
[274]Block A, Dangl J L, Hahlbrock K, et al. Functional borders, genetic fine structure, and distance requirements of cis elements mediating light responsiveness of the parsley chalcone synthase promoter[J]. PNAS.1990,87(14):5387-5391.
[275]Svensson J T, Crosatti C, Campoli C, et al. Transcriptome analysis of cold acclimation in barley Albina and Xantha mutants[J]. Plant Physiology.2006,141(1):257-270.
[276]Simpson S D, Nakashima K, Narusaka Y, et al. Two different novel cis-acting elements of erd 1, a clpA homologous Arabidopsis gene function in induction by dehydration stress and dark-induced senescence[J]. The Plant Journal.2003,33(2):259-270.
[277]Rushmore T H, Morton M R, Pickett C B. The antioxidant responsive element. Activation by oxidative stress and identification of the DNA consensus sequence required for functional activity[J]. Journal of Biological Chemistry.1991,266(18):11632-11639.
[278]Quinn J M, Eriksson M, Moseley J L, et al. Oxygen deficiency responsive gene expression in Chlamydomonas reinhardtii through a copper-sensing signal transduction pathway[J]. Plant Physiology.2002,128(2):463-471.
[279]Quinn J M, Barraco P, Eriksson M, et al. Coordinate copper-and oxygen-responsive Cyc6 and Cpxl expression in Chlamydomonas is mediated by the same element[J]. Journal of Biological Chemistry.2000,275(9):6080-6089.
[280]Saha D, Prasad A, Sujatha T, et al. In silico analysis of the Lateral Organ Junction (loj) gene and promoter of Arabidopsis thaliana[J]. In Silico Biology.2007,7(1):7-19.
[281]Urao T, Yamaguchi-Shinozaki K, Urao S, et al. An Arabidopsis myb homolog is induced by dehydration stress and its gene product binds to the conserved MYB recognition sequence[J]. Plant Cell.1993,5(11):1529-1539.
[282]Zhu Y, Cai X L, Wang Z Y, et al. An interaction between a MYC protein and an EREBP protein is involved in transcriptional regulation of the rice Wx gene[J]. Journal of Biological Chemistry. 2003,278(48):47803-47811.
[283]Kucho K, Yoshioka S, Taniguchi F, et al. Cis-acting elements and DNA-binding proteins involved in CO2-responsive transcriptional activation of Cahl encoding a periplasmic carbonic anhydrase in Chlamydomonas reinhardtii[J]. Plant Physiology.2003,133(2):783-793.
[284]Hamilton D A, Schwarz Y H, Mascarenhas J P. A monocot pollen-specific promoter contains separable pollen-specific and quantitative elements[J]. Plant Molecular Biology.1998,38(4): 663-669.
[285]Inukai Y, Sakamoto T, Ueguchi-Tanaka M, et al. Crown rootlessl, which is essential for crown root formation in rice, is a target of an AUXIN RESPONSE FACTOR in auxin signaling[J]. Plant Cell.2005,17(5):1387-1396.
[286]Harper R M, Stowe-Evans E L, Luesse D R,et al. The NPH4 locus encodes the auxin response factor ARF7, a conditional regulator of differential growth in aerial Arabidopsis tissue[J]. Plant Cell.2000,12(5):757-770.
[287]Liu Z B, Ulmasov T, Shi X, et al. Soybean GH3 promoter contains multiple auxin-inducible elements[J]. Plant Cell.1994,6(5):645-657.
[288]Finkelstein R R, Lynch T J. The Arabidopsis abscisic acid response gene ABI5 encodes a basic leucine zipper transcription factor[J]. Plant Cell.2000,12(4):599-609.
[289]Zhang Z L, Xie Z, Zou X, et al. A rice WRKY gene encodes a transcriptional repressor of the gibberellin signaling pathway in aleurone cells[J]. Plant Physiology.2004,134(4):1500-1513.
[290]Xie Z, Zhang Z L, Zou X, et al. Annotations and functional analyses of the rice WRKY gene superfamily reveal positive and negative regulators of abscisic acid signaling in aleurone cells[J]. Plant Physiology.2005,137(1):176-189.
[291]Nishiuchi T, Shinshi H, Suzuki K. Rapid and transient activation of transcription of the ERF3 gene by wounding in tobacco leaves:possible involvement of NtWRKYs and autorepression[J]. Journal of Biological Chemistry.2004,279(53):55355-55361.
[292]Hinderer W, Petersen M, Seitz H U. Inhibition of flavonoid biosynthesis by gibberellic acid in cell suspension cultures of Daucus carota L[J]. Planta.1984,160(6):544-549.
[293]Finkler A, Kaplan B, Fromm H. Ca2+-responsive cis-elements in plants[J]. Plant Signaling & Behavior.2007,2(1):17-19.
[294]Yanagisawa S, Sheen J. Involvement of maize Dof zinc finger proteins in tissue-specific and light-regulated gene expression[J]. Plant Cell.1998,10(1):75-90.
[295]Schmid J, Doerner P, Clouse S, et al. Developmental and environmental regulation of a bean chalcone synthase promoter in transgenic tobacco[J]. Plant Cell.1990,2(7):619-631.
[296]Kubasek W, Shirley B, McKillop A, et al. Regulation of flavonoid biosynthetic genes in germinating Arabidopsis seedlings[J]. Plant Cell.1992,4(10):1229-1236.