摘要
目的:克隆人色素域解旋酶DNA结合蛋白5(chromodomain helicase DNA-binding protein5,CHD5)基因并构建真核表达载体。方法:应用PCR扩增人CHD5基因的编码区,使用基因重组方法构建真核细胞表达载体peGFPc1-CHD5,实时定量PCR技术检测重组质粒peGFPc1-CHD5转染293T工具细胞后的过表达能力。结果:经过PCR方法,有效扩增了CHD5基因编码区,构建了peGFPc1-CHD5真核细胞表达质粒,测序分析表明所克隆的CHD5基因编码区序列无误。实时定量PCR检测结果表明重组质粒peGFPc1-CHD5能有效地过表达CHD5基因。结论:成功地构建了人CHD5的真核细胞表达载体peGFPc1-CHD5,并在293T工具细胞中实现过表达,为进一步研究奠定了实验基础。
Objective：To construct an expression vector that can express CHD5 gene in eukaryotic. Methods：The human gene CHD5 was amplified from human brain cDNA by PCR. The expression vector peGFPc1-CHD5 was constructed by DNA recombination. The constructed plasmid was transformed into 293T cell by lipofectamine 2000. The expression level of CHD5 mRNA in293T cell was determined by real-time PCR. Results：The human gene CHD5 was effectively amplified. The expression vector peGFPc1-CHD5 was constructed,the DNA sequencing result showed that the constructed plasmid containing CHD5 gene was the same as designed. Real-time PCR analysis showed that the recombinant plasmid peGFPc1-CHD5 could express the CHD5 mRNA in 293T cell. Conclusions：The human gene CHD5 has been successfully cloned and expressed,which could be used as the tool for research on function and mechanism of CHD5 gene.
出处
《现代生物医学进展》
CAS
2009年第21期4045-4047,共3页
Progress in Modern Biomedicine
基金
第二军医大学青年启动基金
