番茄WRKY转录因子基因片段的克隆及序列分析

Cloning and Sequence Analysis of WRKY Transcription Factor Gene Fragment in Tomato

以番茄苗为实验材料,克隆出番茄WRKY转录因子的一个基因片段并对其核酸序列进行分析。根据笔者课题组已经克隆出的番茄特异的WRKY基因片段和番茄EST序列设计特异引物,利用RT-PCR技术获得番茄WRKY转录因子的一个基因片段并连接到pMD19-T载体中,转化DH5α,筛选阳性克隆,用双酶切分析法对其进行鉴定并进一步对核酸序列进行测序分析。结果表明,克隆获得的WRKY转录因子基因片段约为680 bp,用GenBank中的Blastn程序分析表明,其与CaWRKY(AY789641.1)、WIZZ(wizz mRNA)(AB028022.1)的相似性分别为86%和84%。WRKY转录因子在番茄中同样受JA诱导,这为克隆番茄WRKY基因全长,进而为番茄抗病育种奠定基础。

In this study, tomato seedlings were used as gene fragment was cloned and sequenced. The specific experimental materials; a WRKY transcription factor primer was designed according to the cloned WRKY transcription factor gene fragment and tomato EST sequences; a gene fragment was amplified by RT-PCR. The fragment was inserted into the pMD19-T vector, and transformed into E. coli DH5ct.The recombinant plasmid was digested with Pst I and EcoR I, and sequenced. The result showed that the fragment was about 680 bp. Blastn analysis showed thatthe similarity with CaWRKY （AY789641.1） and WIZZ （wizz mRNA） （AB028022.1） were 86% and 84% respectively. WRKY transcription factor in tomato was induced by JA, which laid the foun- dation for cloning the full WRKY gene sequence in tomato and breeding disease-resistant tomato.