2型常染色体显性遗传性多囊肾病致病基因突变研究

Detection of the mutations of autosomal dominant polycystic kidney disease gene 2 in Chinese

目的建立检测2型常染色体显性遗传性多囊肾病致病基因PKD2突变的方法,分析中国汉族人PKD2基因的突变。方法收集临床确诊的中国汉族人常染色体显性遗传性多囊肾病(ADPKD)患者48例,用试剂盒提取外周血白细胞DNA,利用聚合酶链反应-单链构象多态性分析(PCR-SSCP)技术进行突变分析,对异常条带的PCR产物进行核苷酸序列测定,明确突变位点和方式。结果从48例中成功检测到4种突变,包括1种无义突变、1种移码突变、2种错义突变。第1种为外显子5的1249C→T,417位编码氨基酸发生无义突变。第2种为外显子13的第2401位碱基A缺失,造成编码氨基酸移码突变。第3种突变为外显子1的568G→A,编码氨基酸改变为190Ala→Thr;第4种为外显子5的1168G→A,编码氨基酸改变为390Gly→Ser。结论PCR-SSCP技术可用于PKD2的直接基因诊断,并从本组患者中检测到4种突变,丰富了PKD2基因突变谱,为今后开展ADPKD的直接基因诊断、产前诊断和囊肿前诊断提供了一种有用方法。

Objective To develop a method for detecting the mutations in autosomal dominant polycystic kidney disease gene 2(PKD2) and detect the mutations of PKD2 in Chinese. Methods The white blood cell genomic DNA from 48 Chinese ADPKD patients was isolated and amplified by polymerase chain reaction(PCR) using the primers designed according to the PKD2 sequence .The PCR products were analyzed by single strand complement polymorphism(SSCP). The samples of abnormal bands were sequenced. Results Four mutations were identified, including 1 nonsense mutation, 1 deletion mutation and 2 missense mutations. The first one was in exon 5(1249 C→T),where a stop code formed. The second was in exon13(2401 delete A),resulting in the reading frame shift mutation. The third and fourth mutations were in exon1 and exon5(568 G→A,1168G→A), causing the amino acid changes(190Ala→Thr,390Gly→Ser). Conclusions The PCR SSCP method can be used in detecting the mutations of PKD2 and successfully in identifying 4 mutations in PKD2 from 48 ADPKD patients. It is helpful in the molecular diagnosis of ADPKD in advance of the cysts formation and birth.