摘要
目的设计一种将亚硫酸氢化测序PCR和甲基化特异性PCR相结合的巢式PCR技术,即BSMSP技术,对原发性肝细胞癌手术切除组织中p16INK4A甲基化情况进行检测。方法基因组DNA经亚硫酸氢化修饰后,首先通过亚硫酸氢化测序PCR评估模板的质量,然后再分别用甲基化和非甲基化特性PCR分析基因的甲基化状态,并将所得的扩增产物进行测序验证。结果在40例HBV感染的肝细胞癌中,有3例(7.5%)因模板质量原因无法检测,其余37例标本中p16INK4A基因甲基化发生率为78%,测序结果证实了所用方法的可行性。结论BSMSP技术对于肿瘤发生过程中基因甲基化的研究,以及临床诊断方面都具有一定的应用价值。
Objective To detection the methylation of p16INK4A in primary hepatocellular carcinoma, a nested bisulfite sequencing and methylation-specific polymerase chain reaction （BS-MSP）protocol was designed and used. Methods Bisulfite-modified DNA were amplified to evaluate the quality oftemplates with a pair of bisulfite sequencing primers in the first round of PCR, then subjected to methylationassay with corresponding methylation or unmethylation specific PCR primers. Representative PCR productswere sequenced to confirm its correctness. Results 3 of 40 cases （7. 5% ） were failed to assay due to poorquality of templates, and 29 of 37 cases （78%） were detected p16INK4A methylation. Sequencing resultsconfirmed that templates were correctly amplified. Conclusion BS-MSP technique might be valuable formethylation study on carcinogenesis and clinical assay.
出处
《中华检验医学杂志》
CAS
CSCD
北大核心
2006年第3期229-232,共4页
Chinese Journal of Laboratory Medicine
基金
天津市重点科技攻关专项基金资助项目(05YFSJSF02500)
天津市重点基金资助项目(033801011)
关键词
甲基化
肝肿瘤
Methylation
Liver neoplasms
