摘要
目的探讨多重置换扩增(MDA)与激光捕获显微切割(LCM)技术相结合,并应用到胃癌细胞杂合性丢失(LOH)分析等研究的可行性。方法利用LCM技术从胃癌组织冰冻切片中分别获取正常胃黏膜细胞和胃癌细胞,提取基因组DNA后,进行全基因组多重置换扩增。进而,将MDA产物用于聚合酶链反应(PCR)扩增ACE2、TP53和ACTB基因片段,以及微卫星位点的LOH分析。结果位于不同染色体的3个基因片段均得到较好的扩增,而且MDA产物的扩增效率明显高于未经MDA的基因组DNA。另外,MDA产物的LOH分析结果与未经MDA的基因组DNA结果一致。结论MDA与LCM技术相结合,是一种可行的全基因组扩增技术路线,可用于后续的基因组学研究。
Objective To develop a feasible technical procedure for obtaining the purified and sufficient genomic DNA used in further genomic analysis of gastric cancer with laser capture microdissection （LCM） and multiple displacement amplification （MDA）. Methods Normal gastric mucosal cells and gastric cancer ceils were captured from frozen gastric cancer sections by using LCM. Genomic DNA was extracted and MDA was performed. MDA product was used for PCR to amplify ACE2 ,TP53 ,ACTB and loss of heterozygosity （LOH） analysis of D17S1308. Results All of the three genes located in at different chromosomes have been successfully amplified, and the amplification egiciency of MDA product was significantly higher than original LCM DNA. In LOH analysis, the result of MDA product was consistent with that of the original LCM DNA. Conclusion MDA combined with LCM can be used to obtain purified, sufficient genomic DNA used in further genomic analysis of zastric cancer.
出处
《中华实验外科杂志》
CAS
CSCD
北大核心
2007年第11期1350-1352,共3页
Chinese Journal of Experimental Surgery
基金
国家基础研究发展规划资助项目(G1998051203)
国家杰出青年科学基金(30125017)
国家自然科学基金(30370640)
关键词
胃癌
杂合性丢失
基因
Gastric cancer
Loss of heterozygosity
Gene
