基于转录组测序黑老虎角鲨烯合酶基因克隆和生物信息学分析  

Cloning and Bioinformatics Analysis of Squalene Synthase(KcSQS) Gene from Kadsura coccinea(Lem.) A. C. Smith Based on Transcriptome Sequencing

作　　者：王丽君[1] 李艳青 谢舒平 石雨荷 朱敏[2] 童巧珍[1,4] 段礼新 WANG Lijun;LI Yanqing;XIE Shuping;SHI Yuhe;ZHU Min;TONG Qiaozhen;DUAN Lixin(Hunan University of Chinese Medicine,Changsha410208Hunan,China;Guangzhou University of Chinese Medicine,Guangzhou510006Guangdong,China;Nanjing University of Chinese Medicine,Nanjing210000Jiangsu,China;Yueyang Hospital of Traditional Chinese Medicine,Yueyang414000Hunan,China)

机构地区：[1]湖南中医药大学,湖南长沙410208 [2]广州中医药大学,广东广州510006 [3]南京中医药大学,江苏南京210000 [4]湖南省岳阳市中医院,湖南岳阳414000

出　　处：《中药新药与临床药理》2019年第7期870-878,共9页Traditional Chinese Drug Research and Clinical Pharmacology

基　　金：广州市科技计划项目(2018-1002-SF-0437);广州中医药大学2017年度“高水平大学建设”“杏林青年学者”项目(A1-AFD018171Z0311)

摘　　要：目的研究黑老虎[Kadsura coccinea(Lem.)A. C. Smith]转录组特征,克隆三萜生物合成上游途径的关键酶基因角鲨烯合酶(KcSQS),为黑老虎资源利用奠定基础。方法对黑老虎不同部位进行转录组测序分析,根据筛选出的KcSQS候选基因序列设计引物,利用PCR技术进行基因克隆,并借助生物学软件进行生物信息学分析。结果克隆得到一个KcSQS基因,包含全长1 227 bp的开放阅读框(ORF),编码408个氨基酸,预测其蛋白分子量为46.65 kD,定位于细胞质膜,含一段跨膜超螺旋结构。该基因在黑老虎叶片中表达量最高,与同科植物厚朴(Magnolia officinalis Rehd. et Wils.)角鲨烯合酶(MoSQS)蛋白序列相似性为76.77%,预测为角鲨烯合酶。结论该研究对黑老虎植物根、茎和叶进行转录组测序,成功克隆了角鲨烯合酶基因,并对其进行生物信息学分析,可为进一步研究黑老虎三萜类化合物的生物合成提供依据。Objective Studying the transcriptome feature and cloning the gene encoding key upstream enzyme of squalene synthase(SQS)of Kadsura coccinea(Lem.)A.C.Smith.This study will contribute the utility of K.coccinea(Lem.)A.C.Smith resource.Methods Transcriptome sequencing was performed on different organs of K.coccinea(Lem.)A.C.Smith.The primers were designed according to the candidate KcSQS unigene sequences,and the gene was cloned by PCR technology.The biological information of KcSQS was analyzed using biological software.Results The KcSQS gene contains a 1227 bp Open Reading Frame(ORF),encodes 408 amino acids and has a protein molecular weight of 46.65 kD.KcSQS is predicted to locate in the cytoplasmic membrane and contain a transmembrane superhelical structure.The gene highly expresses in the leaves and it has a high similarity(76.77%)with the SQS protein sequence of the same family plant Magnolia officinalis Rehd.et Wils.It is predicted to encode asqualene synthase.Conclusion We sequenced the transcriptome of root,stem and leaves of K.coccinea(Lem.)A.C.Smith,successfully cloned the KcSQS gene and analyzed its bioinformatics,which will provide a basis for further study on its function and biosynthesis of triterpenoids in K.coccinea(Lem.)A.C.Smith plant.

关 键 词：黑老虎 角鲨烯合酶 转录组测序 基因克隆 

分 类 号：R282.71[医药卫生—中药学] Q811.4[医药卫生—中医学]