人卵透明带重组融合基因及其表达载体的构建

Construction of recombination human zona peiiucida3/GST fusion protion gene expression vector

目的构建人卵透明带蛋白3(zonapellucida3,ZP3)/GST蛋白融合基因原核表达载体。方法利用PCR法扩增出去N端的信号肽和C端的跨膜区huZP3成熟蛋白编码基因。先将其克隆到pGEM-T载体,进行序列测定和分析,然后将huZP3核心DNA片断克隆入GST蛋白融合原核表达载体pGEX4T-1内。结果获得一个核苷酸长度约为1000bp的基因,与GenBank(NCBI:M60504)公布的ZP3基因序列有99%的同源性,DNA序列分析发现,有一个氨基酸出现变异。酶切鉴定后证明ZP3基因完整插入GST融合蛋白原核表达载体pGEX4T-1中。结论成功构建出含ZP3基因的GST融合蛋白原核表达载体。

[Objective] To construct GST fusion protion gene expression vector which can express human zona peiiucida in Prokaryotic cells. [Methods] The ZP3 gene was amplified by using PCR. The amplified DNA fragment was cloned into pGEM-T vector for sequencing analysis. The ZP3 gene was then subcloned into fusion protion expression vector pGEX4T-1. [Result] A gene of 1 000 bp was obtained and showed a 99% homology with ZP3 gene sequence published in GenBank （NCBI： M60504）. ZP3 gene was inserted into Prokaryotic expression vector pGEX4T-1 correctly. [Conclusion] Having constructed ZP3 GST protion expression vector successfully.