氯化消毒饮用水浓集物对ras基因的DNA损伤作用

ras Gene Damage Induced by Organic Concentrate of Drinking Water

目的运用依赖随机化末端连接物PCR(RDPCR)技术检测氯化消毒饮用水浓集物对ras基因的DNA损伤作用,探讨其可能致癌作用分子机制。方法采用酚-氯仿-异戊醇法提取人的类淋巴母细胞(TK6)的DNA,采用PCR法制备人类N-ras基因外显子1、2和H-ras基因外显子1、2(以下分别简称为N1、N2、H1、H2)单链探针。将采集的某自来水厂水源水和管网末梢水各150L进行浓集,分别以0.1、0.5、1L/ml的剂量(相当于原水)对TK6细胞进行染毒,提取基因组DNA,再经RDPCR扩增,扩增产物与N1、N2、H1、H2单链探针杂交、显色。结果管网末梢水水样浓集物染毒组DNA经N2特异性引物指导下的RDPCR扩增产物,与N2单链探针杂交,在1L/ml浓度组观察到1条杂交显色条带,在0.1、0.5L/ml浓度组均未观察到杂交条带;经H1、H2、N1特异性引物指导下的RDPCR扩增产物与相应的基因特异性单链探针杂交,在0.1、0.5、1L/ml浓度组均未观察到杂交条带。水源水水样浓集物染毒组DNA经N1、N2、H1、H2特异性引物指导下的RDPCR扩增产物与相应的基因特异性单链探针杂交,在0.1、0.5、1L/ml浓度组均未观察到杂交条带。结论氯化消毒饮用水浓集物对N2具有DNA损伤作用,具有遗传毒性。

Objective To detect the DNA damage of ras gene induced by organic concentrate of drinking water using randomized terminal linker-dependent PCR. Methods The single-stranded probes of exons of rass gene in human were prepared. The TK6 cell was treated with organic concentrate of drinking water, then genomic DNA was amplified by RDPCR and detected by Southern hybridization with the probe. Results The clear hybridized band induced by organic concentrate of tap water could be seen in the position of exon 2 of N-ras gene, but in those of exon 1 of N-ras, exon 1 and 2 of H-ras gene it could not be detected. No hybridized bands induced by organic concentrate of raw water were seen in any position. Conclusion The organic concentrate of drinking water can cause the DNA damage of ras gene and it is considered as a kind of genotoxic substance.