Mutation Detection In PKD1 Gene In Chinese Adpkd Families By Using PCR And DNA Sequencing

Objective: Autosomal dominant polycystic kidney disease (ADPKD) is one of the most common inherited renal diseases. The incidence is 1:1,000. It causes 1/10 of all renal failure. There are at least three different genes: PKD1, PKD2 and Tg737, are involved in ADPKD. Mutations of PKD1 are thought to account for approximately 85% of all mutations in ADPKD. PKD1 has been located in 16p13.e and is made up of 46 exons. The search for mutations is one of the most important steps in understanding the molecular mechanisms underlying ADPKD. But be hindered by both its large size and complicated genomic structure. To date, only about 160 mutations of PKD1 have been described, and the most of the mutations have been reported in Caucasian patients. Only a few of mutations of PKD1 were detected in Chinese. We undertook the study scan for mutations in Chinese population in order to find some features about Chinese patients and made a fandmental for further research.Methods: Forty-three PKD1-affected individuals from thirty-eight unrelated families and fifteen control participate in the study. Fifty-three blood samples and five renal tissues were obtained after receiving informed consent and were in accordance with institutional guidelines. Genomic DNA was isolated using ABCD procedure (Shandong Provincial Hospital Experiment Center). PCR amplification of genomic DNA was performed to generate aimed fragments. All amplified fragments were analyzed by sequencing and searching form genebank to make a comparion.Results: Aimed fragments of exon 12, 14 and 21 were amplified and sequenced by using PCR and DNA sequencing techniques. Then analysised in Genebank. Two novel mutations were detected, including one nonsense mutation (C26017A) and one missesse mutation (A33849G) . In addition, one polymorphism(G26874A) were identified in both ADPKD patients and normal person. The mutation detection rate is 4.7%(2/43) in our study.Conclusion: (1) Two novel pathogenic mutations were detected, including onenonsense mutation(C26017A) and one missense mutation (A33849G) . (2) No hot spots in PKD1 duplicated regions were detected in Chinese patients. (3) In contrast to missense mutations, nonsense mutations cause no more serious symptoms in Chinese ADPKD patients. (4) PCR band with sequencing is the most accuracy method for detecting nucleotide changes (mutations or polymorphisms) in a chosen sequence.