检测PKD2基因多态性生物芯片的建立

Fabrication of oligonucleotide microarray for genotyping PKD2 genetic polymorphisms

目的:PKD2基因在人群中以多种等位基因的形式存在和遗传,其中某些变异型等位基因所编码的多囊蛋白2功能发生改变或缺失,从而引起常染色体显性遗传多囊肾病(ADPKD)的发生。检测PKD2基因的变异情况具有重要的临床诊断意义。方法:针对文献报道的中国人群中检测发现的PKD2变异位点,设计检测探针及引物,并制备基因芯片,构建了基因片段的质粒作为标准模板。结果:通过实验优化,建立了样品荧光标记、基因芯片杂交与检测的条件及分型标准。通过对标准质粒模板和人外周血提取的基因组DNA模板的检测,验证了基因芯片检测结果的重复性和准确性。结论:本实验制备的基因芯片可以准确区分PKD2基因中的9个突变位点的变异情况。

Objective：To diagnosie the genetic variation of PKD2 gene which exists and is inherited in the population with various alleles, some of which encode abnormal protein of polyeistin-2 causing autosomal dominant polyeystie kidney disease （ADPKD）. Methods.Aiming at the mutation sites in PKD2 gene detected in Chinese population, detection probes and priers were designed, and an oligonucleotide mieroarray was fabricated. The plasmids containing the target segments were constructed and used as the standard template. Results： Through experiments, conditions for fluorescent labeling, hybridization and detection were optimized, and genotyping standard was established. The reproducibility and accuracy of the genotyping result using oligonueleotide microarray were tested by detection of the standard plasmid template and the human genomic DNA specimens. Conclusion： It was shown that the oligonueleotide microarray fabricated in this study was able to detect the 9 mutation sites in PKD2 gene correctly.