摘要： 采用基于EST的电子克隆方法，以拟南芥(Arabidopsis thaliana)的尿黑酸叶绿基转移酶基因(Homogentisate phytyltransferase，HPT)cDNA序列为信息探针，对大豆的EST数据库进行同源检索筛选，获得了1777 bp长大豆HPT基因的cDNA序列(GenBank登录号为：AY96421)，经RT-PCR扩增、分子克隆和序列分析验证，表明与电子克隆序列一致；该基因具有完整的开放阅读框架(ORF，173～1408 bp)，推测编码411个氨基酸的蛋白。该蛋白与拟南芥(Arabidopsis thaliana)、苜蓿(Medtcago sativa)、水稻(Oryza satwa)、玉米(Zea mays)、光合集胞蓝细菌(Synechocystis)的序列进行了比较，发现该基因具有保守性。

关键词: 电子克隆, EST数据库, RT-PCR, 尿黑酸叶绿基转移酶, 大豆

Abstract: In silico cloning based on EST sequence is a new method of gene cloning, using Arabidopsis thaliana homogentisate phytyltransferase cDNA sequence as a query probe to blast Glycine max EST database, a soybean HPT gene cDNA of 1777 bp was obtained (GeneBank Accession, AY956421). This sequence was confirmed by RT-PCR, molecular cloning and sequencing, It contained a complete ORF, from 173 bp to 1408 bp, encoded 411 amino acids, and conserved with Arabidopsis thaliana, Medicago sative, Oryza, Zea mays and Synechocystis. The results show that it is an efficient technique to clone new genes by searching EST database with homologous gene of model species.

Key words: In silico cloning, EST database, RT-PCR, Homogentisate phytyltransferase, Soybean

中图分类号: 

• S565.103