玉米sbe2a基因RNAi载体的构建及转化玉米的初步研究

The Primary Study on Construction of RNAi Expression Vector of sbe2a Gene and Maize Transformation

采用PCR技术克隆了玉米淀粉分支酶sbe2a基因的cDNA片段,将其克隆到pMD18-T载体,对重组子进行PCR检测和限制性内切酶分析,并进行序列分析。结果表明:克隆片段长度为562 bp,将该基因的正义和反义片段插入到pCAMBIA1301改造过的载体p35-1301的35S启动子下游,构建高效RNAi表达载体,通过花粉管通道法将其导入玉米自交系"铁7922",经过PCR检测获得了4株转基因株系,初步证明外源基因已整合到玉米基因组中。

cDNA segment of corn starch branching enzymes gene sbe2a was cloned by PCR technique and inserted into pMD18-T vector.The recombinant clone was detected by PCR and the sequence and fragments of restriction enzymes were analyzed.The results showed that the length of the clone sequence was 562 bp.The sense and anti-sense fragments of clone sequence were inserted into downstream of 35S promoter of pCAMBIA1301 to construct over-expression RNAi vector.And then it was transformed into corn inbred lines＂ TIE 7922＂ by Pollen-tube Pathway.It was confirmed primarily that the exogenous gene was integrated into corn genome via PCR analyzing the four transgenic plants.It could improve the content of amylase.