无芒隐子草CsLEA基因超表达载体和反义表达载体构建

Construction of the over and antisense expression vector of CsLEA gene

根据已克隆出的无芒隐子草(Cleistogenes songorica)晚期胚胎发生丰富蛋白基因CsLEA(GenBank序列号为FJ972827)基因和植物表达载体的酶切位点特征,分别将CsLEA基因编码区序列480 bp正向和反向插入pBI121,构建了超表达载体pBI121 35S::CsLEA和反义表达载体pBI121 35S::AS-CsLEA。电击法转入根癌农杆菌GV3101中,并通过花粉管通道法转化拟南芥(Arabidopsis thaliana),经卡那霉素抗性筛选和PCR鉴定,已获得T1代阳性植株。

According to the restriction enzyme sites of plant expression vector and sequence of CsLEA gene from Cleistogenes songorica,over expression vector was recombined by inserting the open reading frame( ORF) sequence to pBI121,and the antisense plant expression vector was constructed by ligating 480 bp sequence of CsLEA gene to pBI121 in the antisense orientation. Moreover,the recombined plasmid was transformed into Agrobacterium tumefaciens GV3101 by electroporation method. Using pollen-tube pathway,the plasmid was transformed into Arabidopsis thaliana and T1 positive plants were obtained by kanamycin resistance selection and PCR confirmation.