应用差异PCR技术研究MTX耐药细胞的遗传学改变

基因扩增是介导细胞产生耐药性的一种普遍性机制。为探讨MTX耐药的小鼠细胞中与耐药机制形成有关的分子遗传学背景，应用差异PCR技术，检测了MTX耐药细胞中DHFR基因的扩增和过表达，并对c-myc及p53基因状态与dhfr扩增之间的相关性进行了探讨。结果表明：dhfr的扩增和过表达直接介导细胞耐药。未检测到c-myc的扩增和p53拷贝数的变化；也未检测到c-myc mRNA水平的改变，但p53基因的mRNA水平明显升高，显示p53可正常表达。这些结果表明dhfr的扩增与这两个侯选基因的状态无关，提示该耐药细胞中可能存在其他的分子遗传学改变允许基因大量扩增。

Study on the Genetic Alterations in MTX-resistant Cells by Differential Polymerase Chain Reaction

Gene amplification is a common mechanism that contributes to the drug resistance.To explore the molecular genetic background related to the MTX resistance in the mouse MTX resistant cells,differential PCR was used to determine the amplification and overexpression of DHFR gene.In addition,the correlations between c myc,p53 status and dhfr amplification were studied.Amplification and overexpression of dhfr suggested its role in MTX resistant cells.However,no amplification and overexpression of c myc were detected.On the other hand,no alteration of p53 copy number was found.The increased mRNA level of p53 suggested the normal function of p53 .These results implicated the status of c myc and p53 had no correlation with dhfr amplification,therefore some other molecular genetic alterations may exist to permit the dhfr amplification in MTX resistant cells.