Zhou Hua-Rong, Shen Jian-Zhen, Fu Hai-Yin, Ye Bao-Guo, Fan Li-Ping, Lin Fu-An
Fujian Institute of Hematology, Union Hospital, Fujian Medical University, Fuzhou 350001, China.
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2006 Apr;14(2):375-8.
This study was aimed to investigate the efficiency of modified methylation-specific polymerase chain reaction i.e. nested methylation-specific polymerase chain reaction, used to detect the promoter methylation of p16 gene in six hematological malignant cell lines, and to explore the application in selection of hematological malignant cell lines with promoter hypermethylation, and make them to be an idel cell models for studying the relationship between gene methylation and expression. DNAs were denatured by NaOH and then were subjected to bisulfite modification and a nested-MSP was used to amplify the promoter region, nested MSP product of p16 gene promoter was analyzed and sequenced. The results showed that the hypermethylation of p16 gene was detected in CA46 and U266, however, Molt4, K562, HL-60 and Jurkat cell lines were unmethylated. In conclusion, p16 gene methylation in hematological malignant cell lines can be perfectly detected by nested-MSP method, which is simple, sensitive and specific for screening all kinds of hematological malignant cell lines with p16 gene methylated.
本研究旨在探讨改良的甲基化特异性聚合酶链反应即巢式甲基化特异性聚合酶链反应，用于检测六种血液系统恶性肿瘤细胞系中p16基因启动子甲基化的效率，探索其在筛选启动子高甲基化血液系统恶性肿瘤细胞系中的应用，并使其成为研究基因甲基化与表达关系的理想细胞模型。DNA经NaOH变性后进行亚硫酸氢盐修饰，采用巢式甲基化特异性聚合酶链反应扩增启动子区域，对p16基因启动子的巢式甲基化特异性聚合酶链反应产物进行分析和测序。结果显示，在CA46和U266中检测到p16基因的高甲基化，而Molt4、K562、HL-60和Jurkat细胞系未甲基化。综上所述，巢式甲基化特异性聚合酶链反应方法可完美检测血液系统恶性肿瘤细胞系中的p16基因甲基化，该方法简单、灵敏且特异，可用于筛选各种p16基因甲基化的血液系统恶性肿瘤细胞系。
