芪合酶基因酵母表达载体的构建及其表达研究
|
摘要
本试验应用基因重组技术构建葡萄芪合酶基因(stilbene synthase,STS)重组酵母菌,通过半乳糖诱导外源基因表达,验证芪合酶基因在酵母菌中的表达情况,进而为开发一种含有白藜芦醇生物活性成分的酵母微生态制剂提供了重要的试验素材。
将从中国野生属葡萄种高抗白粉病的华东葡萄白河-35-1中分离出的芪合酶基因cDNA片段,进行PCR扩增;把纯化回收的目的基因PCR产物定向插入克隆载体pGEM-T Easy中进行测序与序列分析;经DNA测序,克隆片段全长1195 bp,其中编码芪合酶基因的片段长1179 bp(包括起始密码子和终止密码子),共编码392个AA,与Genbank中已有的葡萄芪合酶基因进行同源性比对,其核苷酸同源性达98 %,由此推导出的氨基酸序列与已公布的氨基酸序列同源性为99 %,且位于第164位的半胱氨酸(Cys)酶活中心的碱基未发生突变。
将芪合酶基因与克隆载体pGEM-T Easy连接的重组质粒pGEM-T-STS以及酵母表达载体pYES6/CT用BamH I/Xho I双酶切,采用定向克隆的方法将纯化回收的芪合酶基因克隆到大肠杆菌-酵母菌穿梭型载体pYES6/CT上,转化大肠杆菌,提取阳性重组质粒转化酿酒酵母菌(Saccharomyces cerevisiae)菌株INVSc1,经杀稻瘟素(Bsd)抗性筛选出阳性克隆转化子,并进行菌落PCR鉴定;酵母菌落PCR电泳结果显示,在约1200 bp处出现一条亮带,证明芪合酶基因酵母表达载体pYES6/CT-STS构建成功。
将含有阳性单克隆的酵母菌用半乳糖诱导表达,收集诱导表达4、8、12、16、20及24 h的酵母菌采用超声波破碎菌体,并进行全蛋白SDS-PAGE电泳分析;SDS-PAGE电泳分析发现与空白菌相比含目的基因的诱导菌有目标带(48 kDa)出现,证明目的基因cDNA片段可以在酵母菌中表达。
本试验成功构建了葡萄芪合酶基因酿酒酵母表达载体,并实现了在酿酒酵母菌的大量表达,为下一步利用该菌株生产微生态制剂奠定了基础。
With the technology of gene recombination, the recombinant yeast of STS gene was constructed. It may provide important experiment materials for developing a yeast micro-ecological agent containing resveratrol.
A cDNA fragment of STS gene derived from Vitis pesudoreticulata Baihe-35-1 was amplified by PCR. And then the recycling target gene was inserted into the clone vector pGEM-T Easy and carried on sequence analysis. After sequencing, the length of the cloned fragment was 1195 bp and the fragment encoding stilbene synthase gene was 1179 bp (including the start codon and termination codon) which encode 392 AA. The identity of the gene with the grape STS gene registered in Genbank was 98 % and the derived protein was 99 %. And the enzyme activity center-cysteine (Cys) which was the 164th AA did not mutate at all.
Both pGEM-T-STS and pYES6/CT were digestioned by BamH I and Xho I . With the directional cloning method the reclaimed and purified STS gene was cloned into pYES6/CT, a shuttle type plasmid vector of E.coli-yeast and transformed into E.coli. The positive vector was transformed into Saccharomyces cerevisiae INVSc1 strains. The positive colones were screened out with Bsd and further identified by the colony PCR. The yeast colony PCR results showed that a 1200 bp band proved stilbene synthase gene yeast expression vector was constructed successfully.
The yeast containing positive clones were induced with galactose. 4, 8, 12, 16, 20 and 24-hour collection of ultrasonic broken yeast cells was collected and analyzed by SDS-PAGE. By SDS-PAGE analysis the target protein band (48 kDa) appeared in the induced yeast containing the target gene, which identified the target gene cDNA fragment can express in Saccharomyces cerevisiae.
This experiment successfully constructed Saccharomyces cerevisiae grape stilbene synthase gene expression vector, and realized a lot of target protein expression in Saccharomyces cerevisiae. The results layed the foundation for the forward experiment of producing the yeast micro-ecological agent containing resveratrol
将从中国野生属葡萄种高抗白粉病的华东葡萄白河-35-1中分离出的芪合酶基因cDNA片段,进行PCR扩增;把纯化回收的目的基因PCR产物定向插入克隆载体pGEM-T Easy中进行测序与序列分析;经DNA测序,克隆片段全长1195 bp,其中编码芪合酶基因的片段长1179 bp(包括起始密码子和终止密码子),共编码392个AA,与Genbank中已有的葡萄芪合酶基因进行同源性比对,其核苷酸同源性达98 %,由此推导出的氨基酸序列与已公布的氨基酸序列同源性为99 %,且位于第164位的半胱氨酸(Cys)酶活中心的碱基未发生突变。
将芪合酶基因与克隆载体pGEM-T Easy连接的重组质粒pGEM-T-STS以及酵母表达载体pYES6/CT用BamH I/Xho I双酶切,采用定向克隆的方法将纯化回收的芪合酶基因克隆到大肠杆菌-酵母菌穿梭型载体pYES6/CT上,转化大肠杆菌,提取阳性重组质粒转化酿酒酵母菌(Saccharomyces cerevisiae)菌株INVSc1,经杀稻瘟素(Bsd)抗性筛选出阳性克隆转化子,并进行菌落PCR鉴定;酵母菌落PCR电泳结果显示,在约1200 bp处出现一条亮带,证明芪合酶基因酵母表达载体pYES6/CT-STS构建成功。
将含有阳性单克隆的酵母菌用半乳糖诱导表达,收集诱导表达4、8、12、16、20及24 h的酵母菌采用超声波破碎菌体,并进行全蛋白SDS-PAGE电泳分析;SDS-PAGE电泳分析发现与空白菌相比含目的基因的诱导菌有目标带(48 kDa)出现,证明目的基因cDNA片段可以在酵母菌中表达。
本试验成功构建了葡萄芪合酶基因酿酒酵母表达载体,并实现了在酿酒酵母菌的大量表达,为下一步利用该菌株生产微生态制剂奠定了基础。
With the technology of gene recombination, the recombinant yeast of STS gene was constructed. It may provide important experiment materials for developing a yeast micro-ecological agent containing resveratrol.
A cDNA fragment of STS gene derived from Vitis pesudoreticulata Baihe-35-1 was amplified by PCR. And then the recycling target gene was inserted into the clone vector pGEM-T Easy and carried on sequence analysis. After sequencing, the length of the cloned fragment was 1195 bp and the fragment encoding stilbene synthase gene was 1179 bp (including the start codon and termination codon) which encode 392 AA. The identity of the gene with the grape STS gene registered in Genbank was 98 % and the derived protein was 99 %. And the enzyme activity center-cysteine (Cys) which was the 164th AA did not mutate at all.
Both pGEM-T-STS and pYES6/CT were digestioned by BamH I and Xho I . With the directional cloning method the reclaimed and purified STS gene was cloned into pYES6/CT, a shuttle type plasmid vector of E.coli-yeast and transformed into E.coli. The positive vector was transformed into Saccharomyces cerevisiae INVSc1 strains. The positive colones were screened out with Bsd and further identified by the colony PCR. The yeast colony PCR results showed that a 1200 bp band proved stilbene synthase gene yeast expression vector was constructed successfully.
The yeast containing positive clones were induced with galactose. 4, 8, 12, 16, 20 and 24-hour collection of ultrasonic broken yeast cells was collected and analyzed by SDS-PAGE. By SDS-PAGE analysis the target protein band (48 kDa) appeared in the induced yeast containing the target gene, which identified the target gene cDNA fragment can express in Saccharomyces cerevisiae.
This experiment successfully constructed Saccharomyces cerevisiae grape stilbene synthase gene expression vector, and realized a lot of target protein expression in Saccharomyces cerevisiae. The results layed the foundation for the forward experiment of producing the yeast micro-ecological agent containing resveratrol
引文
[1] Bavaresco, Petegolli D, Gantu E, et al. Elicition and accumulation of stilbene phytoalexins in grapeberries infected by Botrytis cinerea [J] .Vitis, 1997, 36(2): 77-83.
[2] Belguendouzl Fremotl. Resveratrol inhibit metal ino-dependence and independent peroxidation of porcine low-density lipoproteins. Biochemical basis of resistance to Botrytis cinerea in grapeberries [J] ,Eppo Bulletin, 1982, 12(2): 167-170.
[3] Chiron H, Drouet A, Lieutier F, et al. Gene induction of stilbene biosynthesis in Scots pine in response to ozone treatment, wounding, and fungal infection [J] .Plant Physiol 2000, 124: 865-872.
[4] Stein U, Blaich R. Studies on stilbene production and susceptibility to Botrytis in vitis species [J] .Vitis, 1985, 24(2): 75-86.
[5] Stein U, Hoos G. Methods for induction and determination of stilbene in the Vitaceae [J] .Vitis, 1984, 23(3): 179-194.
[6] Hain R, Reif H J, Krause E, et al. Disease resistance results from foreign phytoalexin expression in a novel plant [J] .Nature, 1993, 361: 153-156.
[7] Morita H, Noguchi H, Schroder J, et al. Novel polyketides synthesized with a higher stilbene synthase [J] .Eur J Biochem, 2001, 268: 3759-3766.
[8] Hart H. Rote of phytostilbenes in decay and disease resistance [J] .Ann Rev Phyto-Pathol, 1981, 19: 437-458.
[9] Schoppner A, Kindl H. Purification and properties of a stilbene synthase from Induced cell suspension cultures of peanut [J] .J Biol Chem, 1984, 259: 6806-6811.
[10] Schroder G, Brown JW and Schroder J. Molecular analysis of resveratral synthase: cDNA, genomic clones and relationship with chalone synthase [J] .European Journal of Biochemistry, 1988 172: 161-169.
[11] Raiber S, Schroder G, Schroder J. Molecular and enzymatic characterization of two stilbene synthases from Eastern white pine (Pinus strobes). A single Arg/His difference determine the activity and the pH dependence of the enzymes [J] .FEBS Lett, 1995, 361: 299-302
[12] Ingham JL. 3, 4′, 5-trihydroxy stilbene as a phytoalexin from ground nuts (Arachishy-pogaea) [J] .Phytochemistry, 1976, 15: 1791-1793.
[13] Fligmann J, Chroder G, Schanz S, et al. Molecular analysis of chalcone and dihydropinosylvin synthase from sects pine (Pinus sylvestris), and differential regulation of these and related enzyme-activities in stressed plants [J] .Plant Mol Biol, 1992, 18:489-503.
[14] Tropf S, Karcher B, Schroder G, Schroder J. Reaction mechanism of Homo-dimeric plant polyketide synthase (stilben and chalcon synthase) [J] .Biochem Chem, 1995, 270(14): 7922-7937.
[15] Lanz T, Tropf S, Marner FJ, Schroder G. The role of cysteines in polyketi desynthase [J] .Biochem Chem, 1991, 266(15): 9971-9976.
[16] Schroder G, Schroder J. A single change of histidine to glutamine alters the sub- strate preference of a stilben synthase [J] ,Biochem Chem, 1992, 267(29): 20558-20560.
[17] Preisig-Muller R, Gehler R, Melchior F, Stietz U, Kindl H. Plant polyketide synthases leading to stilbenoids have a domain catalyzing malonyl-CoA: CO2 exchange, malonyl-CoA decarboxylation,and covalent enzyme modification and a site for chain lengthening [J] .Biochemistry, 1997, 36: 8349-8358.
[18] Suh DY, Kagami J, Fukuma K, Sankawa U. Evidence for catalytic cysteine- histidine dyad in chalcone synthase [J] .Biochem Biophys Res Commun, 2000, 275: 725-730.
[19] Fettig S, Hess D.Expression of a chimeric stilbene synthase gene in transgenic wheat lines [J] .Transgenic Research, 1999, 8: 179-189.
[20] 梁 辉 , 郑 近 , 段 霞 瑜 , 等 . 用 基 因 枪 法 获 得 抗 白 粉 病 转 芪 合 酶 基 因 小 麦 [J] . 科 学 通报,1999,44(24):2644-2648.
[21] Thomzik JK, Stenzel K, Stocker R, et al. Synthesis of a grapevine phyto- alexin in transgenic tomatoes (Lycopersicon esculentum Mill) condition resistance against Phytophthora infestans [J] .Physiological and Molecular Plant Pathology, 1997, 51: 265-278.
[22] 谭琳,康有法,马冰刚等,芪合成酶基因转化番茄产生白藜芦醇的研究[J] .生命科学研究,2003,7(3):262-266.
[23] Stark-Lorenzen P, Nelke B, H?n?ler G, et al. Transfer of a grapevine stilbene synthase gene to rice (Oryza sativa L) [J] .Plant Cell Reports, 1997, 16: 668-673.
[24] 郭志鸿,王汉宁,陶玲,等.ubi启动子驱动的花生芪合酶基因表达载体的构建及玉米遗传转化初[J] .甘肃农业大学学报, 2004, 39(2):117-123.
[25] Hain R, Bieseler B, Kindl H, et al. Expression of a stilbene synthase gene in Nicotiana tabacum results in synthesis of the phytoalexin resveratrol [J] .Plant Mol-Biol, 1990, 15: 325-335.
[26] Otsuka-chem. Disease- resistant transformed tobacco plant and its construction by genetic transformation [J]. Derwent biotechnology Abstract, 1993, 12(4): 9302190.
[27] 郝捍东,何立人,赵伟珍.白藜芦醇抗柯萨奇病毒感染性小鼠心肌炎的治疗研究[J] .中成药,2003,25(5):398-401.
[28] 刘 芳 , 曲 极 冰 , 李 红 , 等 . 白 藜 芦 醇 抗 炎 作 用 机 制 的 初 步 研 究 [J] . 中 国 药 学 杂志,2006,41(15):1138-1141.
[29] 莫志贤,邵红霞.白藜芦醇苷体外对过氧化氢导致小鼠肝细胞损伤的保护作用[J] .中国药理学通报,2000,16(5):519-521.
[30] 罗 莉 , 黄 忆 明 , 周 光 宇 . 白 藜 芦 醇 对 去 卵 巢 大 鼠 组 织 的 抗 氧 化 作 用 [J] . 中 国 老 年志,2005,25(3):287-289.
[31] 许丹 , 朱明元 , 罗莉 , 等 . 白藜芦醇对衰老小鼠肝脏抗氧化能力影响 [J] . 中国公共卫生,2006,22(4):467-468.
[32] Kerem, Regev-Shoshani Z, Flaishman G, et al. Resveratrol and two monomethylated stilbenes from Israeli Rumex bucephalophorus and their antioxidant potential [J] .Journal of natural products, 2003, 66(9): 1270-1272.
[33] Juan, Emilia Gonzalez-Pons, Eulalia Munuera, et al. trans-Resveratrol, a Natural Antioxidant from Grapes, Increases Sperm Output in Healthy Rats [J] .Journal of nutrition, 2005, 135(4): 757-760.
[34] Cadenas S, Barja G. Resveratrol ,melatonin ,vitamin E ,and PBN protect against renal oxidative DNA damage induced by the kidney carcinogen KBrO3 [J] .Free Radic Biol Med, 1999, 26(11): 1531-1537.
[35] Sgambato A, Ardito R, Faraglia B, et al. Resveratrol, a natural phenolic compound , inhibits cell proliferation and prevents oxidative DNA damage [J] .Mutat Res, 2001, 496(1-2): 171-180.
[36] 刘兆平,于波,李文仙,等.白藜芦醇的雌激素样作用研究[J] .卫生研究,2002,31(3):188-190.
[37] 肖本熙,黄忆明.白藜芦醇对去卵巢大鼠生殖系统的影响[J] .中国公共卫生,2005,21(8):963-964.
[38] Padilla-Banks E, Jefferson, Newbold RR. The immature mouse is a suitable model for detection of estrogenicity in the uterotropic bioassay [J] .Environ Health Perspect, 2001, 109: 821-826.
[39] 唐明增,洪志哲,李向阳,等.白藜芦醇对小鼠免疫系统调节作用的研究[J] .现代中西医结合杂志,2005,14(18):2381-2382.
[40] 俞瑜,曾耀英,季煜华,等.白藜芦醇对小鼠 T 细胞活化、增殖及细胞因子分泌的影响[J] .细胞与分子免疫学杂志,2005,21(6):697-703.
[41] 董玉,戚好文,刘安.白藜芦醇对小鼠肺泡巨噬细胞释放 IL-8 的影响[J] .陕西医学杂志,2006,35(9):1091-1103.
[42] Schwekendiek A, Pfeffer G, Kindl H. Pine stilbene synthase cDNA, a tool for probing environmental stress [J] .FFBS Lett, 1992, 301:41-44.
[43] Schubert R, Fischer R, Hain R, et al. An ozone-responsive region of the grapevine resveratrol synthase promater differs from the basal pathogen-responsive sequence [J] .Plant Mol Biol, 1997, 34: 417-426.
[44] Sparvoli F, Martin C, Scienza A, et al. Cloning and molecular analysis of structural genes involved in flavonoid and stilbene biosynthesis in grape (Vitis vinifera L) [J] .Plant Mol Biol, 1994, 24: 743-755.
[45] Wiese W, Vomam B, Krause E, et al. Structural organization and differential expression of three stilbene synthase genes located on a 13kb grapevine DNA fragment [J] .Plant Mol Biol, 1994, 26: 667-677.
[46] Leckband G, Lorz H. Transformation and expression of a stilbene synthase gene of Vitis vinifera L. In barley and wheat for increased fungal resistance [J] .Theor Appl Genetics, 1998, 96: 1004-1012.
[47] Iinuma Munekazu, Ohyama Masayoshi, Tanaka Toshiyuki. Six flavono stilbenes and aflavanone in roots of sophora alopecuroide [J]. Phytochemistry, 1995, 38(2): 519-526.
[48] Pascual-Marti MC, Salvador A, Chafer A, Berna A. Supercritical fluid extraction of resveratrol from grapes kinof Vitis vinifera and determinat.in by HPLC [J] .Talanta, 2001, 54(4): 735-740.
[49] Sanders Timothy H, McMichael Robert WJr, Hendrix Keith W. Occurrence of resver a troll in edible peanuts [J] .J Agric Food Chem., 2001, 48(4): 1243-1246.
[50] Sobolev VS, ColeR J., Dorner JW, et al. Purification and liquid chromatographic determination of stilbene phytoalexins in peanuts [J] .J.AOAC Int., 1995, 78: 1177-1182.
[51] Jeandet P, Bessis R, Gautheron B. The production of resveratrol (3, 5, 4.-trihydr oxy stilbene) by grape berries indifferent developmental stages [J] .AmJEnol Vitic, 1991, 42: 41-46.
[52] Siemann EH, Creasy LL. Concentration of the phytoalexin resveratrol in wine [J] .Am.J.Enol.Vitic, 1992, 43: 49-52.
[53] Goldberg DM, Yan J, NGE, et al. A global survey of trans-resveratrol concentrations in commercial wines [J] .Am.J. Enol.Vitic., 1995, 46: 159-165.
[54] 陈雷,韩雅珊. 葡萄不同品种和组织白藜芦醇含量的差异[J] .园艺学报,1999,26(2):12-151.
[55] Eder Reinhard, Wendelin Silvia, Vrhovse k U rsa. Resveratrol contents of grapes and red wines independency on vintage year and harvest date [J] .Mitteil 1ungenK losterneuburg, 2001, 51(2-3): 64-78.
[56] Jeandet Philippe, Bessis Roger, Sbaghi Mohamed, et al. Resveratrol content of wines of different ages: Relationship with fung disease pressure in the vineyard [J] .American Journal of Enology &Viticulture, 1995, 46(1): 1-4.
[57] Vidhyasekaran P, Lalithaku mari D, Govindas wamy CV. Production of a phytoalexin in groundnut dueto storage fungi [J] .Indian Phytopathol, 1972, 25: 240-245.
[58] Dorner JW, Cole RJ, Sanders TH, et al. Interrelationship of kernel water activity, soil temperature, maturity, and phytoalexin production in preharvest aflatoxin contamination of drout-stressed peanuts [J] .Mycopathologia, 1998, 105: 117-128.
[59] Victor S, Sobolev, Richard J, Cole. trans-resveratrol content in commercial peanuts and peanut products [J] .Agric.Food Chem., 1999, 47: 1435-1439.
[60] 朱鸿津.HPLC 测定虎杖中白藜芦醇的含量[J] .中国野生植物资源,2001,20(1):49-501
[61] Cantos E, Espin JC, Tomas-Barberan FA. Postharvest induction modeling method using UV irradiation pulses for obtain ingresveratrol-enriched table grapes: a new functional fruit [J] .Agric Food Chem, 2001, 49(10): 5052-5058.
[62] Gonzalez-CandelasL, Gil JV, Lamuela-Raventos RM, Ramon D. Theuse of transgenic yeasts expressing a gene encod in gaglycosyl-hydrolase as a tool to increase resveratrol content in wine [J] .Int Food Microbiol, 2000, 59(3): 179-183.
[63] Jang M, Cai L, Udeani GO, Slowing KV, et al. Cancer chemopreventive activity of resveratrol,anatural product derived from grapes [J] .Science, 1997, 275: 218-200.
[64] Kotha Subbara maiah, WenJing Chung, Pedro Michaluart, et al. Dannenberg. Resveratrol inhibits cyclo oxygenase- transcription and activity in phorbolester-treated humanm ammary epithelial cells [J] .Biol Chem, 1998, 273: 21875-21882.
[65] Martinez J, Moreno JJ. Effect of resveratrol, a natural polyphenolic compound, on reactive oxygen species and prostagl an production [J] .Biochem Pharmacol, 2000, 59(7): 865-870.
[66] Jang M, Pezzuto JM. Effect of resveratrol on 1, 2-O-tetradecanoyl phorbol-13-acetate-induce doxidative events a gene expression in mouse skin [J] .Cancer Lett, 1998, 134(1): 81-89.
[67] Shih A, Davis FB, Lin HY, Davis PJ. Resveratrol induces a poptosis in thyroid cancercel lines via a MAPK-and p53 dependent mechanism [J] .Clin Endocrinol Metab, 2002, 87(3): 1223-1232.
[68] Ahmad N, Adhami VM, Afaq F, et al. Resveratrol causes WAF- 1/ p21- mediated G (1)-phase arrest of cell cycle and induction of apoptosis in human epidermoid carcinoma A431 cells [J] .Clin Cancer Res, 2001, 7(5): 1466-1473.
[69] Holian O, Walter RJ. Resveratrol inhibits the proliferation of normal human keratinocytes in vitro [J] .J Cell Biochem, 2001, 81(S36): 55-62.
[70] 冯永红,许实波.白藜芦醇药理作用研究进展[J] .国外医药植物药分册, 1996,11(4):155-157.
[71] 闫静,王振月,刘丹宁,刘晓辉.白藜芦醇及其甙的生物活性研究进展[J] .中医药学报, 2000,2:39-40.
[72] 赵霞,陆阳,陈泽乃.白藜芦醇的化学药理研究进展.中草药, 1998,29(12):837-83.
[73] Burkhardt S, Reiter RJ, Tan DX, et al. DNA oxidatively damaged by chromium(11)and H202 is protected by theantioxidants melatonin, N(1)-acetyl-N(2)-fomyl-5-methoxyky nuramine, resveratrol and uric acid [J] .Int J Biochem Cell Biol, 2001, 33(8): 775-783.
[74] Chan MM. Antimicrobial effect of resveratrol on dermatophytes and bacterial pathogens of the skin [J] .Biochem Pharmacol, 2002, 63(2): 99-104.
[75] Frankel EN, Parks EJ, Xu R., et al. Effect of fatty acid-rich fishoil supplement a tionontheoxida tionoflowde sityli poproteins [J] .Lipids, 1994, 29(4): 233-236.
[76] Goldberg DM, Hahn SE, Parkes JG. Beyondal cohol: beverage consumption and cardiovas cularmor tality [J] .Clin Chimica Acta, 1995, 237: 155-187.
[77] Bertelli AA, Giovannini L, Giannessi D, et al. Antiplatelet activity of synthetic and natural resveratrol in red wine [J] .Tissue React, 1995, 17(1): 1-3.
[78] Wang Z, Zou J, Huang Y, et al. Effect of resveratrol on platelet aggregation in vivo and vitro [J] .Chin Med J (Eng), 2002, 115(3): 378-380.
[79] Zou J, Huang Y, Chen Q, et al. Effects of resveratrol on oxidative modification of human low-density lipoprotein [J] .Med J (Engl), 2000, 113(2): 99-102.
[80] Wallerath T, Li H , Godtel-Ambrust U , et al . A blend of poly-phenolic compounds explains the stimulatory effect of red wine on human endot helial NO synthase [J] .Nitric Oxide, 2005, 12 : 972104.
[81] El-Mowafy AM., Abou-Zeid LA, Edafiogho I. Recognition of resveratrol by the human estrogen receptor- alpha: molecula rmodeling approach to under stand its biological actions [J] .Med Princ Pract, 2002, 11(2): 86-92.
[82] Gehm BD, Mcandrews JM, Chien PY, et al. Resveratrol, a polyphenolic compound found in grapes and wine, isanagonist for the estrogen receptor [J] .Proc Natl acad Sci U S A, 1997, 94(25): 14138-14143.
[83] Jennifer L, Bowers, Valentyn V, et al. Resveratrol acts as a mixed agonist/ an-tagonist for estrogen receptors a and b [J] .Endocrinology, 2000, 141(10): 3657-3667.
[84] Bhat KP, Lantvit D, Christov K., et al. Estrogenic and antiestrogenic propertys of resveratrol in mammary tumor models[J].Cancer Res, 2001, 61(20): 7456-7463.
[85] Clack-Walker GD, Miklos GLG. Localization and quantification of circular DNA in yeast [J] .Eur. J. Biochem, 1974, 41: 359-365.
[86] Hinnen A, O'Neill KM, Fink GR. Transformation of yeast [J] .Proceedings of the National Academy of Sciences of the United States of America, 1978, 75(4): 1929-1933.
[87] Hitzeman RA, Hagie FE, Levine HL, et al. Expression of human gene for interferon in yeast [J] .Nature, 1981, 293: 717-722.
[88] Albert H, James BH, Gerald RF. Transformation of yeast [J] .Proc.Natl.Acad.Sci.USA, 1978, 75(4): 1929-1933.
[89] Jean DB. Transformation of yeast by a replicating hybrid plasmid [J] .Nature, 1978, 275(5676): 104-109.
[90] 毛小洪,蔡金科.酵母完整细胞快速高效转化法[J] .生物工程学报,1990,6(2):905-910.
[91] 秦玉静,刘世利,鲍晓明等.高效的酵母完整细胞转化法[J] .生物技术,1997,7(3):37-38.
[92] Ito H, Fukuda Y, Murata K, et al. Transformation of Intact Yeast Cells Treated with Alkali Cations [J] .Journal of Bacteriology, 1983, 153(1): 163-168.
[93] Orr-Weaver TL, Szotak JW, Rothstein RJ. Yeast transformation: a model system for the study of recombination [J] .Proc. Natl. Acad. Sci. USA., 1981, 78(10): 6354-6358.
[94] Sakai Y, Rogi T, Yonehara T, et al. High level ATP production by a genetically-engineered Candida yeast [J] .Biotechnology, 1994, 12: 291-293.
[95] Lopes TS, De Wijs IJ, Steenhauer SI, et al. Factor affecting the miotic stability of high-cope-number integration into the ribosomal DNA of Saccharomyces cerevisiae [J] . yeast, 1996, 12: 467-477.
[96] 罗进贤,何鸣,李文清,等.α-淀粉酶和糖化酶在酿酒酵母中的表达和分泌[J] .生物工程学报,1994,10(4):299-305.
[97] 刘巍峰,高东,王祖农.酿酒酵母(Saccharomyces cerevisiae)半乳糖可诱导表达系统特性的研究[J] .菌物系统,1998,17(3):256-261.
[98] 卢文菊,罗进贤,刘向辉,等.人血管抑素 cDNA 在酿酒酵母中的表达和分泌[J] .广州医学院学报,2000,28(2):7-10.
[99] 兰和魁, 封志纯,黄建生,等.人肺表面活性相关蛋白 A1 在酿酒酵母中表达、纯化及活性分析[J] .生物工程学报,2001,17(4):410-413.
[100] 赵颖怡,梁世中,黄鲲,等.一种新型酿酒酵母附加型分泌表达载体的构建[J] .微生物学报,2002,42(4):431-435.
[101] 于 寒 颖 , 杨 谦 . 核 盘 菌 arom 基 因 的 克 隆 及 其 在 酿 酒 酵 母 中 的 表 达 [J] . 微 生 物 学报,2006,46(1):43-47.
[102] 贾向东,陈喜文,陈德富,等.谷胱甘肽 S-转移酶 Zeta 类基因在酿酒酵母中的表达[J] .遗传,2006,28(5):551-556 .
[103] 李敏,马涛,生举正,等.稳定高效表达木糖还原酶基因工业酿酒酵母的构建及木糖醇发酵的初步研究[J] .食品与发酵工业,2006,32(1):1-5.
[104] Volschenk H, Viljoen M, Grobler J, et al. Engineering a pathway for malate degradation in Saccharomyces cerevisiae [J] .Nature Biotechnol, 1997, 15: 253-257.
[105] Volschenk H, Viljoen, et al. Malolactic fermentation in grape must by a genetically engineered strain of Saccharomyces cerevisiae [J] .Am J Enol Vitic, 1997, 48: 193-196.
[106] Volschenk H, Viljoen M, et al. Malo-ethanolic fermentation in grape must by recombinant strain of Saccharomyces cerevisiae [J] .Yeast, 2001, 18: 963-970.
[107] Labarre C, Guzzo J, Cavin JF, et al. Cloning and characterization of the genes encoding the malolactic enzyme and the malate permease [J] .Applied and Environmental microbiology, 1996, 62(4): 1274-1284.
[108] Mikiko Ikeda, Kimiko Umami, Misato Hinohara, et al. Functional expression of Acetabularia acetabulum vacuolar H+-pyrophosphatase in a yeast VMA3-deficient strain [J] .Journal of Experimental Botany, 2002, 53(378): 2273-2275.
[109] Eksteen JM, Van rensburg P, Cordero Otero RR, et al. Starch fermentation by recombinant saccharomyces cerevisiae strains expressing the alpha-amylase and glucoamylase gene from lipomyces kononenkoae and saccharomycopsis fibuligera [J] .Biotechnol Bioeng, 2003, 84(6): 639-646.
[110] Moraes LM, Astolfi-Filho S, Oliver SG. Development of yeast strains for the efficient utilisation of starch: evaluation of constructs that express alpha-amylase and glucoamylase separately or as bifunctional fusion proteins [J] .Applied Microbiology & Biotechnology, 1995 43, (6): 1067-1076.
[111] Chung Jun-Lan, Sen Alaattin, Wang-Buhler Woon-Gye, et al. cDNA-directed expression of a functional zebrafish CYP1A in yeast [J] .Aquatic Toxicology (Amsterdam), 2004, 70(2): 111-121.
[112] Biswas EE. Heterologous expression of the human retina specific ABC transporter, ABCR, in the yeast Saccharomyces cerevisiae [J] .IOVS, 2004, 45(Suppl. 1): 523.
[113] Lim Chae-Kwon, Kim Kwang-Hyeon, Kim Chul-Ho, et al. Secretion and localization of Pseudomonas auratiaca levansucrase expressed in Saccharomyces cerevisiae [J] .Journal ofMicrobiology & Biotechnology, 2004, 32(3): 206-211.
[114] 陈赞谋,齐章旺.酵母培养物对断奶仔猪的饲喂效果试验[J] .饲料工业,2006,27(15):39-40.
[115] 李洪龙,董艺凝,孙明梅.酵母菌培养物对蛋鸡生产性能及蛋品质的影响[J] .中国饲料,2006,(13):19-20.
[116] Carlson MS, Veum TL, Turk JR. Effects of yeast extract versus animal plasma in weanling pig diets on growth performance and intestinal morphology [J] .Journal of swine health and production, 2005, 13(4): 204-209.
[117] Zhang AW, Lee BD, Lee SK, et al. Effects of yeast (Saccharomyces cerevisiae) cell components on growth performance, meat quality, and ileal mucosa development of broiler chicks [J] .Poultry science, 2005, 84(7): 1015-1021.
[118] 龚月生,任转龙,刘宏斌等.酵母及其培养物对反刍动物生产性能的影响[J] .黄牛杂志,2002,28(5):30-33.
[119] 王聪,任金焕,刘强等.酵母对奶牛泌乳性能及健康状况影响的研究[J] .兽药与饲料添加剂,2005,10(1):7-9.
[120] 陈丽,唐修君,王杏龙.精料中添加酵母培养物对犊牛生长的影响[J] .现代畜牧兽医,2005, (11):16-17.
[121] Wohlt JE, Finkelstein AD, Chung CH. Yeast culture to improve intake, nutrient digestibility, and performance by dairy cattle during early lactation[J] .Journal of dairy science, 1991, 74(4): 1395-1400.
[122] Drennan MJ, Moloney AP. Effect of yeast culture on growth of beef cattle fed on grass silage plus barley-based concentrates [J] .Irish journal of agricultural and food research, 1993, 32(2): 125-132.
[123] Onyango, Bedford EM, Adeola MR. The yeast production system in which Escherichia coli phytase is expressed may affect growth performance, bone ash, and nutrient use in broiler chicks [J] .Poultry science, 2004, 83(3): 421-427.
[124] 黄遵锡,慕跃林,李凤梅,等.基因工程酵母产植酸酶的应用性质研究[J] .饲料研究,2000,(6):11-12.
[125] 祝建洪 , 孙建 , 陈珈 , 等 . 植酸酶分泌型酵母工程菌的构建 [J] . 南开大学学报 ( 自然科版),2002,35(4):1-5.
[126] 马进 , 白俊杰 , 李新辉 , 等 . 虹鳟 生 长激 素 cDNA 在 酵 母 中 的 表 达 [J] . 生 物工 程学报,1999,15(4):434-438.
[127] 陈荣忠,双宝,张锐,等.重组鲈鱼生长激素酵母饲料添加剂及对网箱喂养海水鱼的促长效应[J] .台湾海峡,2003,22(2):181-186.
[128] 温刘发,何丹林,张常明,等.抗菌肽酵母制剂的生产及其作饲料添加剂应用价值的探讨[J] .广东蚕业,2001,35(2):34-36.
[129] 刘进元,常智杰,赵广荣.分子生物学实验指导[M] .北京: 清华大学出版社,2002.
[130] Adams A, Gottschling DE, Kaiser CA, et al 主编. 刘子铎译.Methods in Yeast Genetics: A Cold Spring Harbor Laboratory Course manual[M] .北京: 科学出版社,2000.
[131] Schroeder G, Brown J S, Schroeder J. Molecular analysis fresveratrol synthase: cDNA genomic clones and relationship with chalcone synthase [J] .Eur J Biochem, 1988,172: 161-169.
[132] Goodwin Ph, Hsiang T, Erickson L. A comparison of stilbene and chalcone synthase including a new stilbene synthase gene from Vitis riparia cv [J] . Gloire de Montpellier Plant Sci, 2000,l31(1):1-8.
[133] Cai W, La P. Resveratrol funtion in transgenic plant. Genbank, Accession number: AF274281.
[134] Kobayashi S, Ding C, Nakamura Y, et al. Kiwifiuits (Actinidia deliciosa transformed with a Vitis stilbene synthase gene produce piceid resveratrol-glucoside) [J]. Plant Cell Rep, 2000,(19):904-910.
[135] 顾淑萍,刘正,宋培智.乳杆菌的生长曲线和世代时间测定[J] .上海口腔医学,1998,7(4):226-227.
[136] 任晓燕,剡根强,周霞.粪肠球菌生长曲线的测定[J] .畜牧兽医杂志,2005,24(5):10-11.
[137] Morita H, Noguchi H, Schroder J, et al, Novel polyketides synthesized with a higher plant stilbene synthase [J] . Eur J Biochem, 2001, 268:3759-3766.
[138] 陈惠.花生白藜芦醇合酶基因的克隆及原核表达[D] .福建师范大学:2004.
[139] Melchior F, Kindl H. Grapevine stilbene synthase cDNA only slightly differing from chalcone synthase cDNA is expressed in Escherichia coil into a catalyically active enzyme [J] .FEBS Lett, 1990,268(1):17-20.
[140] 金晓丽.葡萄白藜芦醇合成酶基因的克隆及其在毕赤酵母中的表达[D] .西北大学:2004.
[2] Belguendouzl Fremotl. Resveratrol inhibit metal ino-dependence and independent peroxidation of porcine low-density lipoproteins. Biochemical basis of resistance to Botrytis cinerea in grapeberries [J] ,Eppo Bulletin, 1982, 12(2): 167-170.
[3] Chiron H, Drouet A, Lieutier F, et al. Gene induction of stilbene biosynthesis in Scots pine in response to ozone treatment, wounding, and fungal infection [J] .Plant Physiol 2000, 124: 865-872.
[4] Stein U, Blaich R. Studies on stilbene production and susceptibility to Botrytis in vitis species [J] .Vitis, 1985, 24(2): 75-86.
[5] Stein U, Hoos G. Methods for induction and determination of stilbene in the Vitaceae [J] .Vitis, 1984, 23(3): 179-194.
[6] Hain R, Reif H J, Krause E, et al. Disease resistance results from foreign phytoalexin expression in a novel plant [J] .Nature, 1993, 361: 153-156.
[7] Morita H, Noguchi H, Schroder J, et al. Novel polyketides synthesized with a higher stilbene synthase [J] .Eur J Biochem, 2001, 268: 3759-3766.
[8] Hart H. Rote of phytostilbenes in decay and disease resistance [J] .Ann Rev Phyto-Pathol, 1981, 19: 437-458.
[9] Schoppner A, Kindl H. Purification and properties of a stilbene synthase from Induced cell suspension cultures of peanut [J] .J Biol Chem, 1984, 259: 6806-6811.
[10] Schroder G, Brown JW and Schroder J. Molecular analysis of resveratral synthase: cDNA, genomic clones and relationship with chalone synthase [J] .European Journal of Biochemistry, 1988 172: 161-169.
[11] Raiber S, Schroder G, Schroder J. Molecular and enzymatic characterization of two stilbene synthases from Eastern white pine (Pinus strobes). A single Arg/His difference determine the activity and the pH dependence of the enzymes [J] .FEBS Lett, 1995, 361: 299-302
[12] Ingham JL. 3, 4′, 5-trihydroxy stilbene as a phytoalexin from ground nuts (Arachishy-pogaea) [J] .Phytochemistry, 1976, 15: 1791-1793.
[13] Fligmann J, Chroder G, Schanz S, et al. Molecular analysis of chalcone and dihydropinosylvin synthase from sects pine (Pinus sylvestris), and differential regulation of these and related enzyme-activities in stressed plants [J] .Plant Mol Biol, 1992, 18:489-503.
[14] Tropf S, Karcher B, Schroder G, Schroder J. Reaction mechanism of Homo-dimeric plant polyketide synthase (stilben and chalcon synthase) [J] .Biochem Chem, 1995, 270(14): 7922-7937.
[15] Lanz T, Tropf S, Marner FJ, Schroder G. The role of cysteines in polyketi desynthase [J] .Biochem Chem, 1991, 266(15): 9971-9976.
[16] Schroder G, Schroder J. A single change of histidine to glutamine alters the sub- strate preference of a stilben synthase [J] ,Biochem Chem, 1992, 267(29): 20558-20560.
[17] Preisig-Muller R, Gehler R, Melchior F, Stietz U, Kindl H. Plant polyketide synthases leading to stilbenoids have a domain catalyzing malonyl-CoA: CO2 exchange, malonyl-CoA decarboxylation,and covalent enzyme modification and a site for chain lengthening [J] .Biochemistry, 1997, 36: 8349-8358.
[18] Suh DY, Kagami J, Fukuma K, Sankawa U. Evidence for catalytic cysteine- histidine dyad in chalcone synthase [J] .Biochem Biophys Res Commun, 2000, 275: 725-730.
[19] Fettig S, Hess D.Expression of a chimeric stilbene synthase gene in transgenic wheat lines [J] .Transgenic Research, 1999, 8: 179-189.
[20] 梁 辉 , 郑 近 , 段 霞 瑜 , 等 . 用 基 因 枪 法 获 得 抗 白 粉 病 转 芪 合 酶 基 因 小 麦 [J] . 科 学 通报,1999,44(24):2644-2648.
[21] Thomzik JK, Stenzel K, Stocker R, et al. Synthesis of a grapevine phyto- alexin in transgenic tomatoes (Lycopersicon esculentum Mill) condition resistance against Phytophthora infestans [J] .Physiological and Molecular Plant Pathology, 1997, 51: 265-278.
[22] 谭琳,康有法,马冰刚等,芪合成酶基因转化番茄产生白藜芦醇的研究[J] .生命科学研究,2003,7(3):262-266.
[23] Stark-Lorenzen P, Nelke B, H?n?ler G, et al. Transfer of a grapevine stilbene synthase gene to rice (Oryza sativa L) [J] .Plant Cell Reports, 1997, 16: 668-673.
[24] 郭志鸿,王汉宁,陶玲,等.ubi启动子驱动的花生芪合酶基因表达载体的构建及玉米遗传转化初[J] .甘肃农业大学学报, 2004, 39(2):117-123.
[25] Hain R, Bieseler B, Kindl H, et al. Expression of a stilbene synthase gene in Nicotiana tabacum results in synthesis of the phytoalexin resveratrol [J] .Plant Mol-Biol, 1990, 15: 325-335.
[26] Otsuka-chem. Disease- resistant transformed tobacco plant and its construction by genetic transformation [J]. Derwent biotechnology Abstract, 1993, 12(4): 9302190.
[27] 郝捍东,何立人,赵伟珍.白藜芦醇抗柯萨奇病毒感染性小鼠心肌炎的治疗研究[J] .中成药,2003,25(5):398-401.
[28] 刘 芳 , 曲 极 冰 , 李 红 , 等 . 白 藜 芦 醇 抗 炎 作 用 机 制 的 初 步 研 究 [J] . 中 国 药 学 杂志,2006,41(15):1138-1141.
[29] 莫志贤,邵红霞.白藜芦醇苷体外对过氧化氢导致小鼠肝细胞损伤的保护作用[J] .中国药理学通报,2000,16(5):519-521.
[30] 罗 莉 , 黄 忆 明 , 周 光 宇 . 白 藜 芦 醇 对 去 卵 巢 大 鼠 组 织 的 抗 氧 化 作 用 [J] . 中 国 老 年志,2005,25(3):287-289.
[31] 许丹 , 朱明元 , 罗莉 , 等 . 白藜芦醇对衰老小鼠肝脏抗氧化能力影响 [J] . 中国公共卫生,2006,22(4):467-468.
[32] Kerem, Regev-Shoshani Z, Flaishman G, et al. Resveratrol and two monomethylated stilbenes from Israeli Rumex bucephalophorus and their antioxidant potential [J] .Journal of natural products, 2003, 66(9): 1270-1272.
[33] Juan, Emilia Gonzalez-Pons, Eulalia Munuera, et al. trans-Resveratrol, a Natural Antioxidant from Grapes, Increases Sperm Output in Healthy Rats [J] .Journal of nutrition, 2005, 135(4): 757-760.
[34] Cadenas S, Barja G. Resveratrol ,melatonin ,vitamin E ,and PBN protect against renal oxidative DNA damage induced by the kidney carcinogen KBrO3 [J] .Free Radic Biol Med, 1999, 26(11): 1531-1537.
[35] Sgambato A, Ardito R, Faraglia B, et al. Resveratrol, a natural phenolic compound , inhibits cell proliferation and prevents oxidative DNA damage [J] .Mutat Res, 2001, 496(1-2): 171-180.
[36] 刘兆平,于波,李文仙,等.白藜芦醇的雌激素样作用研究[J] .卫生研究,2002,31(3):188-190.
[37] 肖本熙,黄忆明.白藜芦醇对去卵巢大鼠生殖系统的影响[J] .中国公共卫生,2005,21(8):963-964.
[38] Padilla-Banks E, Jefferson, Newbold RR. The immature mouse is a suitable model for detection of estrogenicity in the uterotropic bioassay [J] .Environ Health Perspect, 2001, 109: 821-826.
[39] 唐明增,洪志哲,李向阳,等.白藜芦醇对小鼠免疫系统调节作用的研究[J] .现代中西医结合杂志,2005,14(18):2381-2382.
[40] 俞瑜,曾耀英,季煜华,等.白藜芦醇对小鼠 T 细胞活化、增殖及细胞因子分泌的影响[J] .细胞与分子免疫学杂志,2005,21(6):697-703.
[41] 董玉,戚好文,刘安.白藜芦醇对小鼠肺泡巨噬细胞释放 IL-8 的影响[J] .陕西医学杂志,2006,35(9):1091-1103.
[42] Schwekendiek A, Pfeffer G, Kindl H. Pine stilbene synthase cDNA, a tool for probing environmental stress [J] .FFBS Lett, 1992, 301:41-44.
[43] Schubert R, Fischer R, Hain R, et al. An ozone-responsive region of the grapevine resveratrol synthase promater differs from the basal pathogen-responsive sequence [J] .Plant Mol Biol, 1997, 34: 417-426.
[44] Sparvoli F, Martin C, Scienza A, et al. Cloning and molecular analysis of structural genes involved in flavonoid and stilbene biosynthesis in grape (Vitis vinifera L) [J] .Plant Mol Biol, 1994, 24: 743-755.
[45] Wiese W, Vomam B, Krause E, et al. Structural organization and differential expression of three stilbene synthase genes located on a 13kb grapevine DNA fragment [J] .Plant Mol Biol, 1994, 26: 667-677.
[46] Leckband G, Lorz H. Transformation and expression of a stilbene synthase gene of Vitis vinifera L. In barley and wheat for increased fungal resistance [J] .Theor Appl Genetics, 1998, 96: 1004-1012.
[47] Iinuma Munekazu, Ohyama Masayoshi, Tanaka Toshiyuki. Six flavono stilbenes and aflavanone in roots of sophora alopecuroide [J]. Phytochemistry, 1995, 38(2): 519-526.
[48] Pascual-Marti MC, Salvador A, Chafer A, Berna A. Supercritical fluid extraction of resveratrol from grapes kinof Vitis vinifera and determinat.in by HPLC [J] .Talanta, 2001, 54(4): 735-740.
[49] Sanders Timothy H, McMichael Robert WJr, Hendrix Keith W. Occurrence of resver a troll in edible peanuts [J] .J Agric Food Chem., 2001, 48(4): 1243-1246.
[50] Sobolev VS, ColeR J., Dorner JW, et al. Purification and liquid chromatographic determination of stilbene phytoalexins in peanuts [J] .J.AOAC Int., 1995, 78: 1177-1182.
[51] Jeandet P, Bessis R, Gautheron B. The production of resveratrol (3, 5, 4.-trihydr oxy stilbene) by grape berries indifferent developmental stages [J] .AmJEnol Vitic, 1991, 42: 41-46.
[52] Siemann EH, Creasy LL. Concentration of the phytoalexin resveratrol in wine [J] .Am.J.Enol.Vitic, 1992, 43: 49-52.
[53] Goldberg DM, Yan J, NGE, et al. A global survey of trans-resveratrol concentrations in commercial wines [J] .Am.J. Enol.Vitic., 1995, 46: 159-165.
[54] 陈雷,韩雅珊. 葡萄不同品种和组织白藜芦醇含量的差异[J] .园艺学报,1999,26(2):12-151.
[55] Eder Reinhard, Wendelin Silvia, Vrhovse k U rsa. Resveratrol contents of grapes and red wines independency on vintage year and harvest date [J] .Mitteil 1ungenK losterneuburg, 2001, 51(2-3): 64-78.
[56] Jeandet Philippe, Bessis Roger, Sbaghi Mohamed, et al. Resveratrol content of wines of different ages: Relationship with fung disease pressure in the vineyard [J] .American Journal of Enology &Viticulture, 1995, 46(1): 1-4.
[57] Vidhyasekaran P, Lalithaku mari D, Govindas wamy CV. Production of a phytoalexin in groundnut dueto storage fungi [J] .Indian Phytopathol, 1972, 25: 240-245.
[58] Dorner JW, Cole RJ, Sanders TH, et al. Interrelationship of kernel water activity, soil temperature, maturity, and phytoalexin production in preharvest aflatoxin contamination of drout-stressed peanuts [J] .Mycopathologia, 1998, 105: 117-128.
[59] Victor S, Sobolev, Richard J, Cole. trans-resveratrol content in commercial peanuts and peanut products [J] .Agric.Food Chem., 1999, 47: 1435-1439.
[60] 朱鸿津.HPLC 测定虎杖中白藜芦醇的含量[J] .中国野生植物资源,2001,20(1):49-501
[61] Cantos E, Espin JC, Tomas-Barberan FA. Postharvest induction modeling method using UV irradiation pulses for obtain ingresveratrol-enriched table grapes: a new functional fruit [J] .Agric Food Chem, 2001, 49(10): 5052-5058.
[62] Gonzalez-CandelasL, Gil JV, Lamuela-Raventos RM, Ramon D. Theuse of transgenic yeasts expressing a gene encod in gaglycosyl-hydrolase as a tool to increase resveratrol content in wine [J] .Int Food Microbiol, 2000, 59(3): 179-183.
[63] Jang M, Cai L, Udeani GO, Slowing KV, et al. Cancer chemopreventive activity of resveratrol,anatural product derived from grapes [J] .Science, 1997, 275: 218-200.
[64] Kotha Subbara maiah, WenJing Chung, Pedro Michaluart, et al. Dannenberg. Resveratrol inhibits cyclo oxygenase- transcription and activity in phorbolester-treated humanm ammary epithelial cells [J] .Biol Chem, 1998, 273: 21875-21882.
[65] Martinez J, Moreno JJ. Effect of resveratrol, a natural polyphenolic compound, on reactive oxygen species and prostagl an production [J] .Biochem Pharmacol, 2000, 59(7): 865-870.
[66] Jang M, Pezzuto JM. Effect of resveratrol on 1, 2-O-tetradecanoyl phorbol-13-acetate-induce doxidative events a gene expression in mouse skin [J] .Cancer Lett, 1998, 134(1): 81-89.
[67] Shih A, Davis FB, Lin HY, Davis PJ. Resveratrol induces a poptosis in thyroid cancercel lines via a MAPK-and p53 dependent mechanism [J] .Clin Endocrinol Metab, 2002, 87(3): 1223-1232.
[68] Ahmad N, Adhami VM, Afaq F, et al. Resveratrol causes WAF- 1/ p21- mediated G (1)-phase arrest of cell cycle and induction of apoptosis in human epidermoid carcinoma A431 cells [J] .Clin Cancer Res, 2001, 7(5): 1466-1473.
[69] Holian O, Walter RJ. Resveratrol inhibits the proliferation of normal human keratinocytes in vitro [J] .J Cell Biochem, 2001, 81(S36): 55-62.
[70] 冯永红,许实波.白藜芦醇药理作用研究进展[J] .国外医药植物药分册, 1996,11(4):155-157.
[71] 闫静,王振月,刘丹宁,刘晓辉.白藜芦醇及其甙的生物活性研究进展[J] .中医药学报, 2000,2:39-40.
[72] 赵霞,陆阳,陈泽乃.白藜芦醇的化学药理研究进展.中草药, 1998,29(12):837-83.
[73] Burkhardt S, Reiter RJ, Tan DX, et al. DNA oxidatively damaged by chromium(11)and H202 is protected by theantioxidants melatonin, N(1)-acetyl-N(2)-fomyl-5-methoxyky nuramine, resveratrol and uric acid [J] .Int J Biochem Cell Biol, 2001, 33(8): 775-783.
[74] Chan MM. Antimicrobial effect of resveratrol on dermatophytes and bacterial pathogens of the skin [J] .Biochem Pharmacol, 2002, 63(2): 99-104.
[75] Frankel EN, Parks EJ, Xu R., et al. Effect of fatty acid-rich fishoil supplement a tionontheoxida tionoflowde sityli poproteins [J] .Lipids, 1994, 29(4): 233-236.
[76] Goldberg DM, Hahn SE, Parkes JG. Beyondal cohol: beverage consumption and cardiovas cularmor tality [J] .Clin Chimica Acta, 1995, 237: 155-187.
[77] Bertelli AA, Giovannini L, Giannessi D, et al. Antiplatelet activity of synthetic and natural resveratrol in red wine [J] .Tissue React, 1995, 17(1): 1-3.
[78] Wang Z, Zou J, Huang Y, et al. Effect of resveratrol on platelet aggregation in vivo and vitro [J] .Chin Med J (Eng), 2002, 115(3): 378-380.
[79] Zou J, Huang Y, Chen Q, et al. Effects of resveratrol on oxidative modification of human low-density lipoprotein [J] .Med J (Engl), 2000, 113(2): 99-102.
[80] Wallerath T, Li H , Godtel-Ambrust U , et al . A blend of poly-phenolic compounds explains the stimulatory effect of red wine on human endot helial NO synthase [J] .Nitric Oxide, 2005, 12 : 972104.
[81] El-Mowafy AM., Abou-Zeid LA, Edafiogho I. Recognition of resveratrol by the human estrogen receptor- alpha: molecula rmodeling approach to under stand its biological actions [J] .Med Princ Pract, 2002, 11(2): 86-92.
[82] Gehm BD, Mcandrews JM, Chien PY, et al. Resveratrol, a polyphenolic compound found in grapes and wine, isanagonist for the estrogen receptor [J] .Proc Natl acad Sci U S A, 1997, 94(25): 14138-14143.
[83] Jennifer L, Bowers, Valentyn V, et al. Resveratrol acts as a mixed agonist/ an-tagonist for estrogen receptors a and b [J] .Endocrinology, 2000, 141(10): 3657-3667.
[84] Bhat KP, Lantvit D, Christov K., et al. Estrogenic and antiestrogenic propertys of resveratrol in mammary tumor models[J].Cancer Res, 2001, 61(20): 7456-7463.
[85] Clack-Walker GD, Miklos GLG. Localization and quantification of circular DNA in yeast [J] .Eur. J. Biochem, 1974, 41: 359-365.
[86] Hinnen A, O'Neill KM, Fink GR. Transformation of yeast [J] .Proceedings of the National Academy of Sciences of the United States of America, 1978, 75(4): 1929-1933.
[87] Hitzeman RA, Hagie FE, Levine HL, et al. Expression of human gene for interferon in yeast [J] .Nature, 1981, 293: 717-722.
[88] Albert H, James BH, Gerald RF. Transformation of yeast [J] .Proc.Natl.Acad.Sci.USA, 1978, 75(4): 1929-1933.
[89] Jean DB. Transformation of yeast by a replicating hybrid plasmid [J] .Nature, 1978, 275(5676): 104-109.
[90] 毛小洪,蔡金科.酵母完整细胞快速高效转化法[J] .生物工程学报,1990,6(2):905-910.
[91] 秦玉静,刘世利,鲍晓明等.高效的酵母完整细胞转化法[J] .生物技术,1997,7(3):37-38.
[92] Ito H, Fukuda Y, Murata K, et al. Transformation of Intact Yeast Cells Treated with Alkali Cations [J] .Journal of Bacteriology, 1983, 153(1): 163-168.
[93] Orr-Weaver TL, Szotak JW, Rothstein RJ. Yeast transformation: a model system for the study of recombination [J] .Proc. Natl. Acad. Sci. USA., 1981, 78(10): 6354-6358.
[94] Sakai Y, Rogi T, Yonehara T, et al. High level ATP production by a genetically-engineered Candida yeast [J] .Biotechnology, 1994, 12: 291-293.
[95] Lopes TS, De Wijs IJ, Steenhauer SI, et al. Factor affecting the miotic stability of high-cope-number integration into the ribosomal DNA of Saccharomyces cerevisiae [J] . yeast, 1996, 12: 467-477.
[96] 罗进贤,何鸣,李文清,等.α-淀粉酶和糖化酶在酿酒酵母中的表达和分泌[J] .生物工程学报,1994,10(4):299-305.
[97] 刘巍峰,高东,王祖农.酿酒酵母(Saccharomyces cerevisiae)半乳糖可诱导表达系统特性的研究[J] .菌物系统,1998,17(3):256-261.
[98] 卢文菊,罗进贤,刘向辉,等.人血管抑素 cDNA 在酿酒酵母中的表达和分泌[J] .广州医学院学报,2000,28(2):7-10.
[99] 兰和魁, 封志纯,黄建生,等.人肺表面活性相关蛋白 A1 在酿酒酵母中表达、纯化及活性分析[J] .生物工程学报,2001,17(4):410-413.
[100] 赵颖怡,梁世中,黄鲲,等.一种新型酿酒酵母附加型分泌表达载体的构建[J] .微生物学报,2002,42(4):431-435.
[101] 于 寒 颖 , 杨 谦 . 核 盘 菌 arom 基 因 的 克 隆 及 其 在 酿 酒 酵 母 中 的 表 达 [J] . 微 生 物 学报,2006,46(1):43-47.
[102] 贾向东,陈喜文,陈德富,等.谷胱甘肽 S-转移酶 Zeta 类基因在酿酒酵母中的表达[J] .遗传,2006,28(5):551-556 .
[103] 李敏,马涛,生举正,等.稳定高效表达木糖还原酶基因工业酿酒酵母的构建及木糖醇发酵的初步研究[J] .食品与发酵工业,2006,32(1):1-5.
[104] Volschenk H, Viljoen M, Grobler J, et al. Engineering a pathway for malate degradation in Saccharomyces cerevisiae [J] .Nature Biotechnol, 1997, 15: 253-257.
[105] Volschenk H, Viljoen, et al. Malolactic fermentation in grape must by a genetically engineered strain of Saccharomyces cerevisiae [J] .Am J Enol Vitic, 1997, 48: 193-196.
[106] Volschenk H, Viljoen M, et al. Malo-ethanolic fermentation in grape must by recombinant strain of Saccharomyces cerevisiae [J] .Yeast, 2001, 18: 963-970.
[107] Labarre C, Guzzo J, Cavin JF, et al. Cloning and characterization of the genes encoding the malolactic enzyme and the malate permease [J] .Applied and Environmental microbiology, 1996, 62(4): 1274-1284.
[108] Mikiko Ikeda, Kimiko Umami, Misato Hinohara, et al. Functional expression of Acetabularia acetabulum vacuolar H+-pyrophosphatase in a yeast VMA3-deficient strain [J] .Journal of Experimental Botany, 2002, 53(378): 2273-2275.
[109] Eksteen JM, Van rensburg P, Cordero Otero RR, et al. Starch fermentation by recombinant saccharomyces cerevisiae strains expressing the alpha-amylase and glucoamylase gene from lipomyces kononenkoae and saccharomycopsis fibuligera [J] .Biotechnol Bioeng, 2003, 84(6): 639-646.
[110] Moraes LM, Astolfi-Filho S, Oliver SG. Development of yeast strains for the efficient utilisation of starch: evaluation of constructs that express alpha-amylase and glucoamylase separately or as bifunctional fusion proteins [J] .Applied Microbiology & Biotechnology, 1995 43, (6): 1067-1076.
[111] Chung Jun-Lan, Sen Alaattin, Wang-Buhler Woon-Gye, et al. cDNA-directed expression of a functional zebrafish CYP1A in yeast [J] .Aquatic Toxicology (Amsterdam), 2004, 70(2): 111-121.
[112] Biswas EE. Heterologous expression of the human retina specific ABC transporter, ABCR, in the yeast Saccharomyces cerevisiae [J] .IOVS, 2004, 45(Suppl. 1): 523.
[113] Lim Chae-Kwon, Kim Kwang-Hyeon, Kim Chul-Ho, et al. Secretion and localization of Pseudomonas auratiaca levansucrase expressed in Saccharomyces cerevisiae [J] .Journal ofMicrobiology & Biotechnology, 2004, 32(3): 206-211.
[114] 陈赞谋,齐章旺.酵母培养物对断奶仔猪的饲喂效果试验[J] .饲料工业,2006,27(15):39-40.
[115] 李洪龙,董艺凝,孙明梅.酵母菌培养物对蛋鸡生产性能及蛋品质的影响[J] .中国饲料,2006,(13):19-20.
[116] Carlson MS, Veum TL, Turk JR. Effects of yeast extract versus animal plasma in weanling pig diets on growth performance and intestinal morphology [J] .Journal of swine health and production, 2005, 13(4): 204-209.
[117] Zhang AW, Lee BD, Lee SK, et al. Effects of yeast (Saccharomyces cerevisiae) cell components on growth performance, meat quality, and ileal mucosa development of broiler chicks [J] .Poultry science, 2005, 84(7): 1015-1021.
[118] 龚月生,任转龙,刘宏斌等.酵母及其培养物对反刍动物生产性能的影响[J] .黄牛杂志,2002,28(5):30-33.
[119] 王聪,任金焕,刘强等.酵母对奶牛泌乳性能及健康状况影响的研究[J] .兽药与饲料添加剂,2005,10(1):7-9.
[120] 陈丽,唐修君,王杏龙.精料中添加酵母培养物对犊牛生长的影响[J] .现代畜牧兽医,2005, (11):16-17.
[121] Wohlt JE, Finkelstein AD, Chung CH. Yeast culture to improve intake, nutrient digestibility, and performance by dairy cattle during early lactation[J] .Journal of dairy science, 1991, 74(4): 1395-1400.
[122] Drennan MJ, Moloney AP. Effect of yeast culture on growth of beef cattle fed on grass silage plus barley-based concentrates [J] .Irish journal of agricultural and food research, 1993, 32(2): 125-132.
[123] Onyango, Bedford EM, Adeola MR. The yeast production system in which Escherichia coli phytase is expressed may affect growth performance, bone ash, and nutrient use in broiler chicks [J] .Poultry science, 2004, 83(3): 421-427.
[124] 黄遵锡,慕跃林,李凤梅,等.基因工程酵母产植酸酶的应用性质研究[J] .饲料研究,2000,(6):11-12.
[125] 祝建洪 , 孙建 , 陈珈 , 等 . 植酸酶分泌型酵母工程菌的构建 [J] . 南开大学学报 ( 自然科版),2002,35(4):1-5.
[126] 马进 , 白俊杰 , 李新辉 , 等 . 虹鳟 生 长激 素 cDNA 在 酵 母 中 的 表 达 [J] . 生 物工 程学报,1999,15(4):434-438.
[127] 陈荣忠,双宝,张锐,等.重组鲈鱼生长激素酵母饲料添加剂及对网箱喂养海水鱼的促长效应[J] .台湾海峡,2003,22(2):181-186.
[128] 温刘发,何丹林,张常明,等.抗菌肽酵母制剂的生产及其作饲料添加剂应用价值的探讨[J] .广东蚕业,2001,35(2):34-36.
[129] 刘进元,常智杰,赵广荣.分子生物学实验指导[M] .北京: 清华大学出版社,2002.
[130] Adams A, Gottschling DE, Kaiser CA, et al 主编. 刘子铎译.Methods in Yeast Genetics: A Cold Spring Harbor Laboratory Course manual[M] .北京: 科学出版社,2000.
[131] Schroeder G, Brown J S, Schroeder J. Molecular analysis fresveratrol synthase: cDNA genomic clones and relationship with chalcone synthase [J] .Eur J Biochem, 1988,172: 161-169.
[132] Goodwin Ph, Hsiang T, Erickson L. A comparison of stilbene and chalcone synthase including a new stilbene synthase gene from Vitis riparia cv [J] . Gloire de Montpellier Plant Sci, 2000,l31(1):1-8.
[133] Cai W, La P. Resveratrol funtion in transgenic plant. Genbank, Accession number: AF274281.
[134] Kobayashi S, Ding C, Nakamura Y, et al. Kiwifiuits (Actinidia deliciosa transformed with a Vitis stilbene synthase gene produce piceid resveratrol-glucoside) [J]. Plant Cell Rep, 2000,(19):904-910.
[135] 顾淑萍,刘正,宋培智.乳杆菌的生长曲线和世代时间测定[J] .上海口腔医学,1998,7(4):226-227.
[136] 任晓燕,剡根强,周霞.粪肠球菌生长曲线的测定[J] .畜牧兽医杂志,2005,24(5):10-11.
[137] Morita H, Noguchi H, Schroder J, et al, Novel polyketides synthesized with a higher plant stilbene synthase [J] . Eur J Biochem, 2001, 268:3759-3766.
[138] 陈惠.花生白藜芦醇合酶基因的克隆及原核表达[D] .福建师范大学:2004.
[139] Melchior F, Kindl H. Grapevine stilbene synthase cDNA only slightly differing from chalcone synthase cDNA is expressed in Escherichia coil into a catalyically active enzyme [J] .FEBS Lett, 1990,268(1):17-20.
[140] 金晓丽.葡萄白藜芦醇合成酶基因的克隆及其在毕赤酵母中的表达[D] .西北大学:2004.