摘要
目的:通过比较变性梯度凝胶电泳(denaturing gradient gel electrophoresis,DGGE)、单链构像多态性(single-strandedconformational polymorphism,SSCP)、限制性酶切检测单核苷酸多态性(single nucleotide polymorphism,SNP)的优缺点,为不同条件下选择最适合的SNP检测方法提供借鉴。方法:DGGE、限制性酶切检测150例大肠癌APEX基因第3外显子,SS-CP检测第5外显子,最后结果经直接测序验证。结果:DGGE检测出17例异常泳动条带;测序结果证实该17例均存在1247A→G的SNP;SSCP检测到57例异常条带;经直接测序证实38例有2197T→G的SNP;PCR-限制性酶切结果与DGGE所测结果完全吻合。结论:DGGE检测结果无假阳性,重复性好,准确率高,而且可判断基因型;SSCP简便易行,但存在假阳性与假阴性,而且不能判断基因型;限制性酶切对SNP位点检测及基因分型均能进行较准确的判断。
Objective: To compare the specialties of DGGE, SSCP and PCR - restriction enzyme digestion in detection of SNP. Methods:The PCR products of 150 specimens of surgical resection of colorectal cancers were screened by DGGE, SSCP, restriction enzyme digestion and sequence. Results: PCR products of exon3 of 17 cases showed aberrant bands in DGGE gel; ex-on5 of 57 cases demonstrated different electrophoresis mobility shifts; direct sequencing identified the existence of 1247A→G in the PCR - amplified fragment of exon3 in all the 17 cases and 2197T→G in the PCR - amplified fragment of exon5 in 37 cases among 57 total cases. The result of PCR - restriction enzyme digestion was identical with that of DGGE. Conclusion: The result of DGGE could be easily repeated, genotype could be judged from the result of DGGE, which is veracity on the whole. SSCP is a convenient method but some pseudo positive and negative results could not be avoided, at the same time genotype could not be judged from the result of SSCP. PCR- restriction enzyme digestion could provide exact information and its genotype about defined SNP.
出处
《中国临床医学》
2004年第2期255-257,共3页
Chinese Journal of Clinical Medicine
基金
国家自然科学基金资助(编号:39870741)
关键词
单核苷酸多态性
检测
变性梯度凝胶电泳
大肠癌
限制性酶切
检测
Single nucleotide polymorphism Denaturing gradient gel electrophoresis Single - stranded conformational polymorphism PCR - restriction enzyme digestion
