论文部分内容阅读
构建一个含1．53kb反义c－myc片段的逆转录病毒载体PAS-c-myc，将其通过Lipofectin导入PA317细胞并经G418筛选获得分泌病毒上清液的阳性克隆。病毒上清波感染血管平滑肌细胞（SMC）经G418筛选获得抗性克区。DNA印迹杂交证实了反义c-myc片段整合于SMC基因组中。RNA印迹杂交证实了反义c－mycRNA在SMC中得到了表达，且显著降低了SMC中c－mycmRNA的水平。蛋白印迹杂交证实了反义c－mycRNA抑制了c－myc蛋白的翻译。提示：c－myc基因表达在SMC增殖中起重要作用。
A retroviral vector PAS-c-myc containing a 1.53kb antisense c-myc fragment was constructed and introduced into PA317 cells by Lipofectin and screened by G418 to obtain positive clones secreting virus supernatant. Virus upper wave infection of vascular smooth muscle cells (SMC) were screened by G418 resistant gram area. Southern blot hybridization confirmed that the antisense c-myc fragment was integrated into the SMC genome. Northern blotting confirmed that antisense c-mycRNA was expressed in SMCs and significantly decreased c-myc mRNA levels in SMCs. Western blotting confirmed that antisense c-mycRNA inhibited the translation of c-myc. Tip: c-myc gene expression plays an important role in SMC proliferation.