| 本文已被：浏览 2223次 下载 266次 |
|
| 长链PCR在中国汉族人多囊肾病1型致病基因突变检测中的应用 |
| 张树忠,梅长林,孙田美,赵海丹,张殿勇,周玉琨,李林,张维莉,吴玉梅,沈学飞 |
|
|
| () |
|
| 摘要: |
| 目的:研究特异性分离中国汉族人多囊肾病1型致病基因(polycystic kidney disease gene 1,PKD1)多拷贝区的方法,以排除同源序列对该基因突变检测的干扰.方法:利用与同源序列间的个别碱基差异设计8对能涵盖PKD1多拷贝区的长链PCR引物,分别对两例健康汉族人基因组DNA进行PCR,其扩增产物通过巢式PCR进行序列测定.结果:通过优化PCR体系,尤其是以高质量基因组DNA为模板,适当提高退火温度,经巢式PCR测序证实扩增产物序列与PKD1多拷贝区一致.结论:该研究采用的长链PCR体系可克服同源序列的影响,适用于中国汉族人PKD1多拷贝区的特异性扩增,为进一步通过单链构象多态性分析检测汉族人PKD1突变位点奠定基础. |
| 关键词: 肾,多囊,常染色体显性、聚合酶链反应、DNA突变分析、中国 |
| DOI： |
|
| 基金项目: |
|
| Long-range PCR in detecting mutation of polycystic kidney disease gene 1 in Hans |
| 张树忠,梅长林,孙田美,赵海丹,张殿勇,周玉琨,李林,张维莉,吴玉梅,沈学飞 |
| () |
| Abstract: |
| To specifically amplify the polycystic kidney disease gene 1(PKD1) via long-range PCR. Methods: Eight pairs of primers were designed based on the differences between the PKD1 and its homologues,and the long-range PCR were performed on the DNA of 2 healthy Hans, and the products were sequenced. Results: With optimum conditions of PCR(including good quality genomic DNA as templates, the higher temperature as Tm,and so on).Subsequently,we succeeded in amplifying the 34 exons of the PKD1 multicopy areas and testify them by sequence. Conclusion: The above long-range PCR not only amplified the multicopy areas of the PKD1 distinctively by eliminating the interferes of its homologues,but also help to detect the PKD1 mutations of the Hans via single strand conformation polymorphism. |
| Key words: kidney,polycystic, autosomal dominant polymerase chain reaction DNA mutation analysis China |
