期刊论文

中文

临床检验杂志

1001-764X

2018

36

2

139-141,147

10.13602/j.cnki.jcls.2018.02.17

湖南省卫计委科研计划课题

本校为第一机构

目的构建包含与宫颈癌相关的EGFR基因G719S和T790M位点重组pMD19-exon18-exon20载体,为制备突变型EGFR基因重组载体提供模板。方法以健康人全血基因组DNA作为模板,设计2对有重叠互补区的特异性引物,对EGFR基因的exon18和exon20两片段分别进行扩增,用重叠PCR技术将exon18和exon20片段进行连接,再将连接产物exon18-exon20片段插入pMD19-T质粒,构建成野生型pMD19-exon18-exon20载体,转入至大肠埃希菌感受态细胞DH5α中进行表达,利用菌液PCR和基因组测序进行鉴定。结果琼脂糖凝胶电泳结果显示, exon18和exon20两位点扩增片段在778 bp和596 bp处有清晰的目的条带,两片段的融合产物exon18-exon20片段可...

Objective To construct the recombinant p MD19-exon18-exon20 plasmid containing locus G719 S and T790 M of EGFR gene associated with cervical cancer,which may provide a template for preparing the mutant recombinant vector of EGFR gene. Methods Using the healthy human genome DNA as templates,the segments of exon 18 and exon 20 of EGFR gene were amplified by two pairs of specific primers which were designed based on the sequences of overlapping and complementary area. The amplified segments were linked by overlap PCR. The products of linked exon18...