PLUNC基因启动子区荧光素酶报告载体的构建与鉴定

目的构建人PLUNC（palate，lung and nasal epithelium clone）基因启动子区C-1888T SNP位点不同单倍型荧光素酶报告基因表达载体。方法利用PCR技术得到1888位点不同基因型的PLUNC基因启动子区目的片段．将其分别插入到pMD18-T载体和pGL3-enhancer荧光素酶报告基因载体中并测序。结果成功构建了人类PLUNC基因C-1888T SNP位点上的2种不同纯合子基因型（CC型和TT型）的荧光素酶报告载体（pGL3-enhancer—T型和pGL3-enhancer—C型），且测序得以证实。结论成功构建出的荧光素酶报告载体，为研究PLUNC基因不同的1888SNF位点能否调控不同的PLUNC基因表达提供了基本实验条件。

The Construction of Luciferase Reporter Gene Vectors Containing Different Haplotypes DNA of Human PLUNC Gene Promoter Region

Objective To Construct the luciferase reporter gene vectors containing two different haplotypes on C- 1888T SNP site of human PLUNC gene promoter region. Method The 280bp DNA fragments of PLUNC promoter region including the 1888 SNP site were obtained by polymerase chain reaction （PCR） based on human genome DNA from the subjects with the CC/TT genotypes, which were TA cloned to pMD18- T simple vectors and then were digested by restriction endonucleases Xho Ⅰ and HindⅢ. After fragment recovery, those fragments were ligated to pGL3-enhancer vectors, which were digested by restriction endonucleases Xho Ⅰ and Hind Ⅲ as well. All recombinant plasmids were identified by sequencing. Result The two recombinant plasmids （pGL3-enhancer-T and pGL3-enhancer-C） containing different haplotypes DNA of human PLUNC promoter region were obtained. Conclusion These expression vectors are the important tool for exploring the possibility of the relation between the 1888 SNP and the expression of PLUNC gene.