分子信标实时PCR检测瘢痕相关p53基因SNP方法的建立

Establishment of molecular beacon real-time PCR for detecting p53 gene single nucleotide polymorphism

目的建立分子信标实时PCR检测病理性瘢痕相关p53基因第72密码子单核苷酸多态性的新方法。方法采集瘢痕疙瘩患者外周血28例,提取DNA并用分子信标PCR检测p53基因多态性。设计一对p53基因第72密码子单核苷酸多态性荧光分子信标探针,荧光定量PCR检测该位点单核苷酸(SNP),并与反向斑点杂交法,DNA直接测序等方法比较。结果定量荧光PCR荧光分子信标检测的结果与测序结果吻合率达到100%,高于反向斑点杂交法且简便快捷。结论分子信标实时PCR检测技术适合临床p53基因第72密码子单核苷酸多态性快速诊断。

Objective To establish a method based on molecular beacon real-time PCR for detecting single nucleotide polymorphisms（SNP） in codon 72 of scar-related p53 gene.Methods Two fluorescence-labeled molecular beacon probes were synthesized targeting CCC/CGC SNP of p53 codon 72.The genomic DNA was extracted from the peripheral blood of 28 patients with keloid,and the CCC/CGC SNP of P53 gene codon 72 were assayed with molecular beacon real-time PCR.The results of SNP typing were compared with the results of reverse dot hybridization and confirmed by direct DNA sequencing.Results The goodness of fit of this method was 100% in comparison with direct DNA sequencing,higher than that of reverse dot hybridization.Conclusion Molecular beacon real-time PCR is suitable for rapid clinical detection of SNPs in p53 gene.