通过PCR从人基因组扩增β珠蛋白核基质结合区(matrix attachment region,MAR)及β干扰素MAR,正向及反向克隆至pCAT3载体SV40启动子的上游,分别检测瞬时及稳定表达的情况下MAR在NIH3T3细胞内对CAT报告基因的影响情况.结果显示:瞬时表达情况下,反向及正向插入的MAR均不能提高CAT基因的表达;稳定整合的情况下,插入的β珠蛋白MAR可使CAT报告基因表达水平提高8倍,β干扰素MAR提高3倍,反向及正向插入的MAR没有明显的差别.这表明MAR能在一定程度上提高外源基因的表达水平,并且不同的MAR对外源基因表达的影响存在差异,MAR的插入方向对外源基因的表达水平没有明显的作用.
The β-globin matrix attachment region (MAR) and β-interferon MAR DNA fragments were amplified through PCR, and cloned into the upstream of SV40 promoter of pCAT3 vector in forward and reverse orientation, the CAR enzyme activities of different constructs were examined under transient and stable expression respectively. The results showed that β-globin MAR could increase the CAT gene expression 8 fold, β-interferon MAR 3 fold in stable integration transformants, however there were little effects of MAR on CAT gene expression levels under transient expression. The orientation of two MARs inserted into the vector had no effect on CAT gene expression.
Chinese Journal of Cell Biology