癌变·畸变·突变Issue(4)：247-251,284,6.DOI:10.3969/j.issn.1004-616x.2013.04.001
全基因组扩增技术对DNA甲基化保真性的影响
The effect of whole genomic amplification on DNA methylation fidelity
摘要
Abstract
OBJECTIVE: Pyrosequencing was applied to study the effect of whole genome amplification (WGA) on the fidelity of DNA methylation. METHODS:EpiTect Whole Bisulfitome Kit was used to perform DNA amplification after whole genome bisulfite modification of L02,A549 and HCT116 cells. The DNA methylation of three specific genes (MGMT,GSTP1,P16) and LINE 1 sequence were analyzed by pyrosequencing. RESULTS:The DNA amplification obtained was 246 folds higher in DNA content and > 1 000 bp fragment length which met the basic requirements of DNA methylation detection. The DNA methylation level of LINE1 did not change after WGA,indicating that WGA had no effect on the whole genome methylation. However,the change of specific DNA methylation after WGA correlated with their original methylation level. When the original methylation rate was higher than 60%,the methylation condition of three genes remained stable after WGA. There was significant change with irregular trend of DNA methylation level after WGA when the initial DNA methylation ranged between ≥20%-60%. DNA methylation rate of three genes declined significantly in all cells when the initial DNA methylation level was lower than 20%. In addition,pyrosequencing result showed that the effect of WGA on genes CpG sites methylation level was similar to that on its mean methylation rate. CONCLUSION:WGA technology was suitable for the study of whole genome methylation and specific gene with higher methylation rate (≥60%). However,we must take caution for using the WGA technology when the rate of DNA methylation was relative low (<60%).关键词
DNA甲基化/全基因组扩增/焦磷酸测序/保真性Key words
DNA methylation/whole genome amplification/pyrosequencing/fidelity分类
生物科学引用本文复制引用
王二曼,何志妮,李道传,张标,陈雯,曾晓雯..全基因组扩增技术对DNA甲基化保真性的影响[J].癌变·畸变·突变,2013,(4):247-251,284,6.基金项目
国家杰出青年基金(30925029)，中国博士后科学基金特别资助项目(201104373) （30925029）
