PCR扩增肠癌组织p53基因失败的原因分析与解决

分析与解决PCR扩增肠癌组织p53基因时失败的问题,为如何解决PCR技术在科研应用中的问题提供一个实例.方法 经初步分析,将失败原因确定为DNA模板有问题、采用下列方法处理模板再进行PCR反应：用70%乙醇再清洗模板、用新标本重新提取模板、用PCR反应成功与失败的模板配成混合模板.比较PCR成功与失败所用模板的差异,寻找与证实失败的原因.结果 最终发现溶解DNA模板的TE缓冲液浓度过高,使PCR扩增失败,排除之后获得成功.结论 当PCR扩增失败时,不妨从最简单的反应体系的离子环境先找原因.

THE DETECTION OF P53 GENE IN HUMAN CERVICAL CARCINOMA WITH AND WITHOUT HUMAN PAPILLOMAVIRUS INFECTION

The special primers Of p53 exon 7 as wed as HPV16 E6 and E7 ORFs (Opening Rending Frame) were used with PCR, PCR-SSCP technique, and 35 specimens of cervical carcinoma were examined. The results were as follows: ① HPV16 E6, E7 DNA was found in 25/35 specimens (71. 4%),which proved again HPV16 Infection an important event in cervical carcinogenesis. However only 11/35 (31.42% ) bad E6 and E7 ORFs simultaneously, 3/35 (8. 57%) and 11/35 (31. 42% ) had only E6 or E7 respectively. ② No mutation and LOH (Loss of Heterozygote) of p53 exon 7 were found in allof 35 specimens. Additionally in the present study, we developed a non-isotopic PCR-SSCP method.

肺癌患者P^(53)基因突变的研究

作者应用多聚酶链反应（PCR）结合限制性内切酶位点多态性及单链构象多态性（SSCP）DNA序列分析技术，对20例肺癌患者的肿瘤组织及正常肺组织P53基因进行了研究。结果显示：15例患者的肿瘤组织中，P53基因存在各种类型的基因突变，为P53基因突变在肺癌的致病机制提供了一定的参考资料，并探讨了P53基因突变与肺癌患者的临床分期及病理组织学分型之间的相关性。