Chelex100法从贮存食管拉网脱落细胞涂片中提取DNA并作PCR分析的应用和意义

Utilizing Chelex100 to extract DNA from stored esophageal balloon cells smears for use with PCR.

目的 探讨从贮存 2 0年的食管拉网脱落细胞涂片的微量细胞中抽提DNA的方法和可行性 ;比较p5 3基因在正常食管上皮和重度不典型增生上皮中的突变差异。方法 利用螯合型离子交换树脂Chelex10 0作为介质 ,一步法从食管拉网脱落细胞中提取用作PCR分析的DNA ,共提取食管正常上皮与重度不典型增生上皮细胞涂片各 2 4例 ,采用PCR SSCP分析法进行p5 3基因第 7外显子的突变检测。结果 所有 48例标本中微量细胞的DNA抽提均获成功 ;正常食管组p5 3基因第 7外显子均未发生突变 ,重度不典型增生组 5例发生突变 ,其中 3例在 10年、12年、14年后转变为食管癌。结论 Chelex10 0法简便、高效 ,能从微量细胞中抽提DNA ,使对 2 0年前食管拉网细胞涂片进行分子水平的检测成为可能 ;p5 3基因第 7外显子的突变使食管上皮重度不典型增生细胞具有了癌变趋势。

Objective 1) To investigate the feasibility of extrac t DNA from esophageal balloon cells sme ars which had been stored for 20 years.2 )To compare the mutation of p53 betwee n normal esophageal epicyte and severe d ysplasia.Methods Chelex100 is a chela ting resin,by utilizing Chelex as a medi um.DNA was extracted with a one-step pro cedure from 48 esophageal balloon cells smears,7th exon of p53 gene mutation w as analyzed by single-strand conformatio n polymorphism method(PCR-SSCP).Results 1) DNA were success extracted from 48 specimens.2)Mutation w as occurred on 7th exon in 5 cases.no mutation was detected in normal esophag eal epithelia;In the 5 cases,3 cases occ urred carcinogenesis after 10,12,14 year s.Conclusion 1) Chelex100 is s imple,efficiency.By it,DNA can extracted from trace cells.It is a successful app roach for the gene study of esophageal b alloon cells smears.2) Carcinogenesis do minates in the precancerous lesions dut to 7th exon mutation of p53 gene.