濒危珍稀植物半枫荷的转录组分析
|
摘要
为加强中国特有濒危植物半枫荷资源的保护与利用工作,采用高通量测序平台Illumina HiSeq 2500对其进行转录组测序,将得到的数据过滤后进行de novo组装并聚类去冗余,获得77 629个Unigenes,通过九大功能数据库比对、分析、注释,最终有45 293个Unigenes获得注释信息;其中在KOG按功能分为25个子类,获得25 253个功能注释信息; GO功能注释可分为细胞组分、生物学过程、分子功能3大类、分别可细分为22、26、17亚类(共65个亚类);与KEGG数据库对比,共发现286条代谢通路,其中发现可能与半枫荷药用活性成分相关的次生代谢产物生物合成的177条途径;根据组装结果预测出88个基因家族共1 547个编码转录因子的Unigenes,发现控制药效合成的转录因子家族。另外,根据注释结果检测到12 579个SNP多态位点和预测出57 671个CDS位点。本研究首次对半枫荷转录组进行分析,为深入开展半枫荷分子生物学研究提供基础数据来源。
In order to strengthen the conservation and utilization of China's endangered plant,Semiliquidambar cathayensis,a high-throughput sequencing platform, Illumina HiSeq 2500, was used to sequence the transcriptome of the plants. The obtained data was filtered and assembled by de novo. Then,77 629 unigenes were obtained through classification and redundancy removal. By comparison analysis,annotation of the 9 functional databases,45 293 unigenes get annotation information,which was divided into 25 sub-categories by function in KOG with 25 253 functional annotation information obtained. The functional annotation of GO could be divided into such three parts as cell components,biological processes,and molecular functions,which is composed of 2 226,and 17 subclasses respectively(65 subclasses in total). With the comparison of KEGG database,286 metabolic pathways are found,among which there are 177 pathways to biosynthesis of secondary metabolites,which may be related to the medicinal active components of S. cathayensis. Based on the assembling result,1 547 unigenes encoding the transcription factors are predicted in 88 gene families. Thus,a family of transcription factors controlling pharmacokinetic synthesis are discovered. Based on the annotation results of S. cathayensis,12 579 SNP polymorphic sites are detected and 57 671 CDS sites are predicted. This study is the first time to analyze the transcriptional sequences of S. cathayensis,providing a basic data for the indepth study on molecular biology in S. cathayensis.
In order to strengthen the conservation and utilization of China's endangered plant,Semiliquidambar cathayensis,a high-throughput sequencing platform, Illumina HiSeq 2500, was used to sequence the transcriptome of the plants. The obtained data was filtered and assembled by de novo. Then,77 629 unigenes were obtained through classification and redundancy removal. By comparison analysis,annotation of the 9 functional databases,45 293 unigenes get annotation information,which was divided into 25 sub-categories by function in KOG with 25 253 functional annotation information obtained. The functional annotation of GO could be divided into such three parts as cell components,biological processes,and molecular functions,which is composed of 2 226,and 17 subclasses respectively(65 subclasses in total). With the comparison of KEGG database,286 metabolic pathways are found,among which there are 177 pathways to biosynthesis of secondary metabolites,which may be related to the medicinal active components of S. cathayensis. Based on the assembling result,1 547 unigenes encoding the transcription factors are predicted in 88 gene families. Thus,a family of transcription factors controlling pharmacokinetic synthesis are discovered. Based on the annotation results of S. cathayensis,12 579 SNP polymorphic sites are detected and 57 671 CDS sites are predicted. This study is the first time to analyze the transcriptional sequences of S. cathayensis,providing a basic data for the indepth study on molecular biology in S. cathayensis.
引文
1.Lander E S,Linton L M,Birren B,et al.Initial sequencing and analysis of the human genome[J].Nature,2001,409(6822):860-921.
2.Zeng S H,Xiao G,Guo J,et al.Development of a ESTdataset and characterization of EST-SSRs in a traditional Chinese medicinal plant,Epimedium sagittatum(Sieb.et Zucc.)Maxim[J].BMC Genomics,2010,11(1):94.
3.Barrero R A,Chapman B,Yang Y F,et al.De novo assembly of Euphorbia fischeriana root transcriptome identifies prostratin pathway related genes[J].BMC Genomics,2011,12(1):600.
4.Xiang L,Li Y,Zhu Y J,et al.Transcriptome analysis of the Ophiocordyceps sinensis fruiting body reveals putative genes involved in fruiting body development and cordycepin biosynthesis[J].Genomics,2014,103(1):154-159.
5.Guo X,LI Y,LI C F,et al.Analysis of the Dendrobium officinale transcriptome reveals putative alkaloid biosynthetic genes and genetic markers[J].Gene,2013,527(1):131-138.
6.于永福.中国野生植物保护工作的里程碑---《国家重点保护野生植物名录(第一批)》出台[J].植物杂志,1999,(5):3.Yu Y F.National key protected wild plants list(first batch)[J].Plants,1999,(5):3.
7.环境保护部,中国科学院.中国生物多样性红色名录高等植物卷[M].北京:环境保护部,中国科学院,2013.Ministry of Environmental Protection,Chinese Academy of Sciences.China biodiversity red list higher plant volumes[J].Beijing:Ministry of Environmental Protection,Chinese Academy of Sciences,2013.
8.傅立国.中国植物红皮书:稀有濒危植物[M].北京:科学出版社,1991.Fu L G.Red book of Chinese plants:rare and endangered plants[M].Beijing:Science Press,1991.
9.赵厚涛,宋培浪,韩国营,等.国家Ⅱ级保护植物半枫荷的最新研究进展[J].北方园艺,2010(21):210-212.Zhao H T,Song P L,Han G Y,et al.The latest research progress on Semiliquidambar cathayensis H.T.Chang[J].Northern Horticulture,2010(21):210-212.
10.林华.半枫荷药材质量标准的规范化研究[D].广州:广州中医药大学,2008.Lin H.Study on standardization in medicinal quality criteria of Pterospermum heterophyllum[D].Guangzhou:Guangzhou University of Chinese Medicine,2008.
11.张颖,贠喆,牛蓓,等.药用半枫荷植物资源研究[J].中国农学通报,2008,24(8):432-434.Zhang Y,Yun Z,Niu B,et al.Stadies on the Resources of Medicinal Plants Ban Fenghe[J].Chinese Agricultural Science Bulletin,2008,24(8):432-434.
12.《全国中草药汇编》编写组.全国中草药汇编.上册[M].北京:人民卫生出版社,1975.Compilation Group of the National Compilation of Chinese Herbal Medicines.National compilation of Chinese herbal medicine(first volume)[M].Beijing:People’s Medical Publishing House,1975.
13.胡君,赵中国,刘庆,等.四川省国家Ⅱ级保护植物新记录---半枫荷[J].四川林业科技,2018,39(1):63-65.Hu J,Zhao Z G,Liu Q,et al.A new record of the national secondary protection species---Semiliquidambar cathayensis H.T.Chang in Sichuan Province[J].Journal of Sichuan Forestry Science and Technology,2018,39(1):63-65.
14.廖娜,韦柳斌,金旺,等.金缕半枫荷多酚提取及其抗氧化抗菌活性研究[J/OL].广西植物.[2018-11-12].http://kns.cnki.net/kcms/detail/45.1134.Q.20180522.1744.008.html.Liao N,Wei L B,Jin W,et al.Extraction technology and the antioxidant and antibacterial activities of polyphenols from Semiliquidambar cathayensis[J/OL].Guihaia.[2018-11-12].http://kns.cnki.net/kcms/detail/45.1134.Q.20180522.1744.008.html.
15.周光雄,杨永春,石建功,等.金缕半枫荷化学成分研究[J].中草药,2002,33(7):589-591.Zhou G X,Yang Y C,Shi J G,et al.Studies on chemical constituents from Semiliquidambar cathayensis[J].Chinese Traditional and Herbal Drugs,2002,33(7):589-591.
16.孙静,郑雪凌,崔向珍,等.半枫荷抗乙型肝炎病毒的药理活性研究[J].时珍国医国药,2014,25(10):2391-2393.Sun J,Zheng X L,Cui X Z,et al.A study of inhibition effects on hepatitis B virus of Semiliquidambar cathayensis in vitro[J].Lishizhen Medicine and Materia Medica Research,2014,25(10):2391-2393.
17.谢金兰,王运昌,范剑明.浅谈半枫荷的资源开发和保护[J].农技服务,2007,24(12):85.Xie J L,Wang Y C,Fan J M.On the development and protection of Semiliquidambar cathayensis resources[J].Serves of Agricultural Technology,2007,24(12):85.
18.Haas B J,Papanicolaou A,Yassour M,et al.De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis[J].Nature Protocols,2013,8(8):1494-1512.
19.Moriya Y,Itoh M,Okuda S,et al.KAAS:an automatic genome annotation and pathway reconstruction server[J].Nucleic Acids Research,2007,35(S2):W182-W185.
20.Rice P,Longden I,Bleasby A.EMBOSS:the European molecular biology open software suite[J].Trends in Genetics,2000,16(6):276-277.
21.Mistry J,Finn R D,Eddy S R,et al.Challenges in homology search:HMMER3 and convergent evolution of coiledcoil regions[J].Nucleic Acids Research,2013,41(12):e121.
22.Quinlan A R,Hall I M.BEDTools:a flexible suite of utilities for comparing genomic features[J].Bioinformatics,2010,26(6):841-842.
23.袁灿,彭芳,杨泽茂,等.川芎转录组SSR分析与EST-SSR标记的开发[J].中国中药杂志,2017,42(17):3332-3340.Yuan C,Peng F,Yang Z M,et al.EST-SSR identification,markers development of Ligusticum chuanxiong based on Ligusticum chuanxiong transcriptome sequences[J].China Journal of Chinese Materia Medica,2017,42(17):3332-3340.
24.Garg R,Patel R K,Tyagi A K,et al.De novo assembly of chickpea transcriptome using short reads for gene discovery and marker identification[J].DNA Research,2011,18(1):53-63.
25.Patel S,Shah D B,Panchal H J.De novo RNA seq assembly and annotation of Trigonella foenum-graecum L.(SRR066197)[J].Legume Genomics and Genetics,2014,5(7):1-7.
26.李翠婷,张广辉,马春花,等.野三七转录组中SSR位点信息分析及其多态性研究[J].中草药,2014,45(10):1468-1472.Li C T,Zhang G H,Ma C H,et al.Analysis on SSR loci information in transcriptome of Panax vienamensis var.fuscidiscus and its polymorphism[J].Chinese Traditional and Herbal Drugs,2014,45(10):1468-1472.
27.王森,张震,姜倪皓,等.半夏转录组中的SSR位点信息分析[J].中药材,2014,37(9):1566-1569.Wang S,Zhang Z,Jiang N H,et al.SSR information in transcriptome of Pinellia ternata[J].Journal of Chinese Medicinal Materials,2014,37(9):1566-1569.
28.王东,曹玲亚,高建平.党参转录组中SSR位点信息分析[J].中草药,2014,45(16):2390-2394.Wang D,Cao L Y,Gao J P.Data mining of simple sequence repeats in Codonopsis pilosula transcriptome[J].Chinese Traditional and Herbal Drugs,2014,45(16):2390-2394.
29.魏秀清,许玲,章希娟,等.莲雾转录组SSR信息分析及其分子标记开发[J].园艺学报,2018,45(3):541-551.Wei X Q,Xu L,Zhang X J,et al.Analysis on SSR information in transcriptome and development of molecular markers in wax apple[J].Acta Horticulturae Sinica,2018,45(3):541-551.
30.Jiang X M,Wu Y F,Xiao F M,et al.Transcriptome analysis for leaves of five chemical types in Cinnamomum camphora[J].Hereditas,2014,36(1):58-68.
31.张少平,洪建基,邱珊莲,等.紫背天葵高通量转录组测序分析[J].园艺学报,2016,43(5):935-946.Zhang S P,Hong J J,Qiu S L,et al.Sequencing and analysis of the transcriptome of Gynura bicolor[J].Acta Horticulturae Sinica,2016,43(5):935-946.
32.郑纪伟.柳树转录组高通量测序及SSR标记开发研究[D].南京:南京林业大学,2013.Zheng J W.Research on transcriptome sequencing and SSRmarkers development of Salix[D].Nanjing:Nanjing Forestry University,2013.
33.蔡年辉,邓丽丽,许玉兰,等.基于高通量测序的云南松转录组分析[J].植物研究,2016,36(1):75-83.Cai N H,Deng L L,Xu Y L,et al.Transcriptome analysis for Pinus yunnanensis based on high throughput sequencing[J].Bulletin of Botanical Research,2016,36(1):75-83.
34.赵丽娟,易小娅,曾幼玲.植物逆境相关C2H2型锌指蛋白的研究进展[J].分子植物育种,2016(3):578-585.Zhao L J,Yi X Y,Zeng Y L.Research progress on plant stress related C2H2type zinc finger protein[J].Molecular Plant Breeding,2016(3):578-585.
35.杜玮南,孙红霞,方福德.单核苷酸多态性的研究进展[J].中国医学科学院学报,2000,22(4):392-394.Du W N,Sun H X,Fang F D.The research development of single nucleotide polymorphism[J].Acta Academiae Medicinae Sinicae,2000,22(4):392-394.
2.Zeng S H,Xiao G,Guo J,et al.Development of a ESTdataset and characterization of EST-SSRs in a traditional Chinese medicinal plant,Epimedium sagittatum(Sieb.et Zucc.)Maxim[J].BMC Genomics,2010,11(1):94.
3.Barrero R A,Chapman B,Yang Y F,et al.De novo assembly of Euphorbia fischeriana root transcriptome identifies prostratin pathway related genes[J].BMC Genomics,2011,12(1):600.
4.Xiang L,Li Y,Zhu Y J,et al.Transcriptome analysis of the Ophiocordyceps sinensis fruiting body reveals putative genes involved in fruiting body development and cordycepin biosynthesis[J].Genomics,2014,103(1):154-159.
5.Guo X,LI Y,LI C F,et al.Analysis of the Dendrobium officinale transcriptome reveals putative alkaloid biosynthetic genes and genetic markers[J].Gene,2013,527(1):131-138.
6.于永福.中国野生植物保护工作的里程碑---《国家重点保护野生植物名录(第一批)》出台[J].植物杂志,1999,(5):3.Yu Y F.National key protected wild plants list(first batch)[J].Plants,1999,(5):3.
7.环境保护部,中国科学院.中国生物多样性红色名录高等植物卷[M].北京:环境保护部,中国科学院,2013.Ministry of Environmental Protection,Chinese Academy of Sciences.China biodiversity red list higher plant volumes[J].Beijing:Ministry of Environmental Protection,Chinese Academy of Sciences,2013.
8.傅立国.中国植物红皮书:稀有濒危植物[M].北京:科学出版社,1991.Fu L G.Red book of Chinese plants:rare and endangered plants[M].Beijing:Science Press,1991.
9.赵厚涛,宋培浪,韩国营,等.国家Ⅱ级保护植物半枫荷的最新研究进展[J].北方园艺,2010(21):210-212.Zhao H T,Song P L,Han G Y,et al.The latest research progress on Semiliquidambar cathayensis H.T.Chang[J].Northern Horticulture,2010(21):210-212.
10.林华.半枫荷药材质量标准的规范化研究[D].广州:广州中医药大学,2008.Lin H.Study on standardization in medicinal quality criteria of Pterospermum heterophyllum[D].Guangzhou:Guangzhou University of Chinese Medicine,2008.
11.张颖,贠喆,牛蓓,等.药用半枫荷植物资源研究[J].中国农学通报,2008,24(8):432-434.Zhang Y,Yun Z,Niu B,et al.Stadies on the Resources of Medicinal Plants Ban Fenghe[J].Chinese Agricultural Science Bulletin,2008,24(8):432-434.
12.《全国中草药汇编》编写组.全国中草药汇编.上册[M].北京:人民卫生出版社,1975.Compilation Group of the National Compilation of Chinese Herbal Medicines.National compilation of Chinese herbal medicine(first volume)[M].Beijing:People’s Medical Publishing House,1975.
13.胡君,赵中国,刘庆,等.四川省国家Ⅱ级保护植物新记录---半枫荷[J].四川林业科技,2018,39(1):63-65.Hu J,Zhao Z G,Liu Q,et al.A new record of the national secondary protection species---Semiliquidambar cathayensis H.T.Chang in Sichuan Province[J].Journal of Sichuan Forestry Science and Technology,2018,39(1):63-65.
14.廖娜,韦柳斌,金旺,等.金缕半枫荷多酚提取及其抗氧化抗菌活性研究[J/OL].广西植物.[2018-11-12].http://kns.cnki.net/kcms/detail/45.1134.Q.20180522.1744.008.html.Liao N,Wei L B,Jin W,et al.Extraction technology and the antioxidant and antibacterial activities of polyphenols from Semiliquidambar cathayensis[J/OL].Guihaia.[2018-11-12].http://kns.cnki.net/kcms/detail/45.1134.Q.20180522.1744.008.html.
15.周光雄,杨永春,石建功,等.金缕半枫荷化学成分研究[J].中草药,2002,33(7):589-591.Zhou G X,Yang Y C,Shi J G,et al.Studies on chemical constituents from Semiliquidambar cathayensis[J].Chinese Traditional and Herbal Drugs,2002,33(7):589-591.
16.孙静,郑雪凌,崔向珍,等.半枫荷抗乙型肝炎病毒的药理活性研究[J].时珍国医国药,2014,25(10):2391-2393.Sun J,Zheng X L,Cui X Z,et al.A study of inhibition effects on hepatitis B virus of Semiliquidambar cathayensis in vitro[J].Lishizhen Medicine and Materia Medica Research,2014,25(10):2391-2393.
17.谢金兰,王运昌,范剑明.浅谈半枫荷的资源开发和保护[J].农技服务,2007,24(12):85.Xie J L,Wang Y C,Fan J M.On the development and protection of Semiliquidambar cathayensis resources[J].Serves of Agricultural Technology,2007,24(12):85.
18.Haas B J,Papanicolaou A,Yassour M,et al.De novo transcript sequence reconstruction from RNA-seq using the Trinity platform for reference generation and analysis[J].Nature Protocols,2013,8(8):1494-1512.
19.Moriya Y,Itoh M,Okuda S,et al.KAAS:an automatic genome annotation and pathway reconstruction server[J].Nucleic Acids Research,2007,35(S2):W182-W185.
20.Rice P,Longden I,Bleasby A.EMBOSS:the European molecular biology open software suite[J].Trends in Genetics,2000,16(6):276-277.
21.Mistry J,Finn R D,Eddy S R,et al.Challenges in homology search:HMMER3 and convergent evolution of coiledcoil regions[J].Nucleic Acids Research,2013,41(12):e121.
22.Quinlan A R,Hall I M.BEDTools:a flexible suite of utilities for comparing genomic features[J].Bioinformatics,2010,26(6):841-842.
23.袁灿,彭芳,杨泽茂,等.川芎转录组SSR分析与EST-SSR标记的开发[J].中国中药杂志,2017,42(17):3332-3340.Yuan C,Peng F,Yang Z M,et al.EST-SSR identification,markers development of Ligusticum chuanxiong based on Ligusticum chuanxiong transcriptome sequences[J].China Journal of Chinese Materia Medica,2017,42(17):3332-3340.
24.Garg R,Patel R K,Tyagi A K,et al.De novo assembly of chickpea transcriptome using short reads for gene discovery and marker identification[J].DNA Research,2011,18(1):53-63.
25.Patel S,Shah D B,Panchal H J.De novo RNA seq assembly and annotation of Trigonella foenum-graecum L.(SRR066197)[J].Legume Genomics and Genetics,2014,5(7):1-7.
26.李翠婷,张广辉,马春花,等.野三七转录组中SSR位点信息分析及其多态性研究[J].中草药,2014,45(10):1468-1472.Li C T,Zhang G H,Ma C H,et al.Analysis on SSR loci information in transcriptome of Panax vienamensis var.fuscidiscus and its polymorphism[J].Chinese Traditional and Herbal Drugs,2014,45(10):1468-1472.
27.王森,张震,姜倪皓,等.半夏转录组中的SSR位点信息分析[J].中药材,2014,37(9):1566-1569.Wang S,Zhang Z,Jiang N H,et al.SSR information in transcriptome of Pinellia ternata[J].Journal of Chinese Medicinal Materials,2014,37(9):1566-1569.
28.王东,曹玲亚,高建平.党参转录组中SSR位点信息分析[J].中草药,2014,45(16):2390-2394.Wang D,Cao L Y,Gao J P.Data mining of simple sequence repeats in Codonopsis pilosula transcriptome[J].Chinese Traditional and Herbal Drugs,2014,45(16):2390-2394.
29.魏秀清,许玲,章希娟,等.莲雾转录组SSR信息分析及其分子标记开发[J].园艺学报,2018,45(3):541-551.Wei X Q,Xu L,Zhang X J,et al.Analysis on SSR information in transcriptome and development of molecular markers in wax apple[J].Acta Horticulturae Sinica,2018,45(3):541-551.
30.Jiang X M,Wu Y F,Xiao F M,et al.Transcriptome analysis for leaves of five chemical types in Cinnamomum camphora[J].Hereditas,2014,36(1):58-68.
31.张少平,洪建基,邱珊莲,等.紫背天葵高通量转录组测序分析[J].园艺学报,2016,43(5):935-946.Zhang S P,Hong J J,Qiu S L,et al.Sequencing and analysis of the transcriptome of Gynura bicolor[J].Acta Horticulturae Sinica,2016,43(5):935-946.
32.郑纪伟.柳树转录组高通量测序及SSR标记开发研究[D].南京:南京林业大学,2013.Zheng J W.Research on transcriptome sequencing and SSRmarkers development of Salix[D].Nanjing:Nanjing Forestry University,2013.
33.蔡年辉,邓丽丽,许玉兰,等.基于高通量测序的云南松转录组分析[J].植物研究,2016,36(1):75-83.Cai N H,Deng L L,Xu Y L,et al.Transcriptome analysis for Pinus yunnanensis based on high throughput sequencing[J].Bulletin of Botanical Research,2016,36(1):75-83.
34.赵丽娟,易小娅,曾幼玲.植物逆境相关C2H2型锌指蛋白的研究进展[J].分子植物育种,2016(3):578-585.Zhao L J,Yi X Y,Zeng Y L.Research progress on plant stress related C2H2type zinc finger protein[J].Molecular Plant Breeding,2016(3):578-585.
35.杜玮南,孙红霞,方福德.单核苷酸多态性的研究进展[J].中国医学科学院学报,2000,22(4):392-394.Du W N,Sun H X,Fang F D.The research development of single nucleotide polymorphism[J].Acta Academiae Medicinae Sinicae,2000,22(4):392-394.