人血管内皮细胞生长因子受体sFlt-1在婴儿双歧杆菌中的表达和鉴定

Expression and Characterization of Human Vascular Endothelial Growth Factor Receptor sFlt-1 in Bifidobacterium Infantis

目的:构建重组质粒pET32a-sFlt-1,并将其导入婴儿双歧杆菌,为婴儿双歧杆菌介导的sFlt-1基因转运系统在体内的抗血管生成作用奠定基础。方法:将编码血管内皮细胞生长因子受体sFlt-1胞外区1-3 loop 316个氨基酸残基的cDNA插入到pET32a构建重组表达质粒pET32a-sFlt-1,转化婴儿双歧杆菌,阳性克隆菌经质粒提取、酶切鉴定,得到两条大小分别与基因sFlt-1及pET32a质粒片段大小相符的电泳条带,RT-PCR检测到外源性sFlt-1基因已成功导入婴儿双歧杆菌。结果:经酶切鉴定质粒pET32a-sFlt-1构建成功,转化入婴儿双歧杆菌,并证实其在mRNA水平可表达。结论:成功构建重组质粒pET32a-sFlt-1成功导入婴儿双歧杆菌,并证实其在mRNA水平可表达。

Objective：We have constructed the system pET32a-sFlt-1 and transfected Bifidobacterium Infantis.This reseach would have a valuable contribution to the study on anti-angiogenesis effect of sFlt-1 mediated by Bifidobacterium Infantis.Methods：By inserting VEGF receptor sFlt-1（1-3 loop） cDNA into pET32a,and We have constructed the system pET32a-sFlt-1 and transfected Bifidobacterium Infantis.The plasmid extracted from positive clones was divided into two parts which were similar in size to sFlt-1 gene and pET32a plasmid after digestion of restriction endonuclease.RT-PCR identification showed that foreign sFlt-1 gene had been integrated into positive colony,and its mRNA expression was tested positive.Results：By digestion of restriction endonuclease,we attest to pET32a-sFlt-1 be transfected Bifidobacterium Infantis,the mRNA expression of sFlt-1 was tested positive.Conclusion：We have constructed the system pET32a-sFlt-1 and transfected Bifidobacterium Infantis.RT-PCR identification showed the mRNA expression of sFlt-1 was tested positive.