摘要
目的介绍一种CpG岛甲基化分析方法,即甲基化特异PCR法(methylation-specific PCR,MSP)以及对该方法的改良。方法用亚硫酸氢盐修饰被测DNA后,进行甲基化与非甲基化特异PCR的扩增,并对MSP法进行改良。应用该法分析人宫颈癌Hela细胞中RASSF1A、p16基因5'CpG岛甲基化状态。结果发现人宫颈癌Hela细胞中有RASSF1A、p16基因的甲基化,改良后的MSP更容易获得较好的效果。结论MSP法是一种较为简便、灵敏和具特异性的甲基化检测方法,改良后该方法更加简便可行。
【Objective】 To introduce a new method for studies on methylation status of CpG island：methylation specific polymerase chain reaction（MSP） with some modification. 【Method】 Target DNA was modified by sodium bisulfite, converting all unmethylated, but not methylated, cytodines to uracil, and subsequent amplification with primers specific for methylated versus unmethylation DNA. The status of 5＇CpG island of tumor suppressor gcn eRASSF1A, p16 in human cervical carcinoma Hela cell was analyzed by modified MSP assy. 【Results】 To get good single strand DNA, the authors denatured the target DNA by heating instead of alkaline treatment, modified single strand DNA by 5 mol/L sodium bisulfite for 8～10 hours instead of 3 mol/l sodium bisulfite for 16～18 h, The 5＇CpG island of RASSF1A, p16 gene was methylated in human cervical carcinoma Hela cell. 【Conclusion】 MSP is simple, sensitive, and specific method for detecting the methylation status of any CpG-rieh region and the modifications make it more simple to perform.
出处
《中国现代医学杂志》
CAS
CSCD
北大核心
2006年第2期191-194,共4页
China Journal of Modern Medicine
基金
湖北省自然科学基金资助(2002AB141)
