一个参与N-甲基-N′-硝基-N-亚硝基胍诱发的遗传不稳定的cDNA片段的分离筛选

Isolation of a cDNA fragment participating N-methyl-N′-nitro-N-nitrosoguanidine induced genetic instability in vero cell

为研究甲基硝基亚硝基胍 ( MNNG)诱发猴肾vero细胞遗传不稳定的机理 ,采用 m RNA差异显示分离到的 MNNG处理后表达发生改变的表达序列标识 ( EST)相关基因进行功能筛选 .利用反义核酸阻断技术阻断其相应基因的表达 ,以筛选分离参与非定标性突变形成的 c DNA片段 .现筛选到 1 0号片段 ,其相关基因的表达被阻断后 ,MNNG诱发的 vero细胞非定标突变率下降至自发突变水平 ,提示 1 0号片段相关的基因是非定标突变发生的正相关基因 .

In order to study the mechanism of genetic instability in monkey kidney vero cells induced by N methyl N′ nitro N nitrosoguanidine (MNNG), we screened the function of relevant genes of expressed sequence tags (ESTs) which had altered expression after MNNG exposure and were isolated by means of mRNA differential display in our lab. Through antisense RNA technique, we blocked expression of the relevant gene of fragment and isolated the cDNA fragment related with genetic instability in vero cell induced by MNNG. When expression of fragment 10 was blocked through transcripting antisense RNA, MNNG induced nontargeted mutation frequency decreased to the level of spontaneous mutation. The result reveals that the relevant gene of fragment 10 is positive correlating gene of nontargeted mutagenesis.