菘蓝APX基因克隆及序列分析

为进一步研究菘蓝APX基因的功能,构建了菘蓝APX基因真核表达载体。从菘蓝植株中提取总RNA,逆转录为cDNA,根据APX基因在该类植物中的同源性设计简并引物,利用PCR方法钓取目的基因,将目的基因与T载体连接,PCR检测阳性克隆,同时菌液送往测序公司进行测序。结果表明:测序片段的生物信息学分析证实了该序列与GenBank登录的APX基因一致。说明成功构建了菘蓝APX重组质粒和克隆鉴定了菘蓝APX基因,可进一步用于基因表达和表达产物的功能研究。

Cloningand identification of APX gene fromIsatis indigotica

To construct the recombinant eukaryotic expression plasmid of pTZ57R /T /APX.APXgene was amplified from the genomic DNA of Isatis indigotica by polymerase chain reaction(PCR).APXgene was then inserted into pTZ57R /T eukaryote expression vector.The positive colonies were screened and identified by PCR and sequencing.The specific gene APXwhich was about 1 257bp,was successfully amplified and pTZ57R /T-APX were constructed.The DNA sequence of APXgene was as the same as the APX gene nucleotide sequences published in Genebank.pTZ57R /T-APX recombinant was successfully constructed.And the construction provides the basis for the further study.