建立基于SNaPshot技术的21-羟化酶相关基因CYP21A2突变的快速检测方法  

The establishment of the rapid detection method of 21-hydroxylase gene CYP21A2 mutations based Snapshot technology

作　　者：刘英华[1] 李海波[1] 王本敬[1] 陈瑛[1] 

机构地区：[1]南京医科大学附属苏州医院生殖与遗传中心,苏州215002

出　　处：《中国优生与遗传杂志》2014年第10期24-26,F0002,共4页Chinese Journal of Birth Health & Heredity

基　　金：江苏省"十二五"科教兴卫工程医学重点人才资助项目(RC2011036)

摘　　要：目的建立一种先天性肾上腺皮质增生症21-羟化酶缺陷的快速基因检测方法。方法选择中国人群21-羟化酶缺乏症相关基因CYP21A2的突变热点Exon3 Del 8bp、Q318X、R356W、IVS2 A/C>G、I172N、P30L和V281L共7个位点进行巢式PCR。设计上述突变热点的单链延伸引物进行扩增,采用SNaPshot技术对上述7个突变热点进行一次性基因分型检测。结果建立了一套满足质量控制标准的CYP21A2基因快速检测的方法,5例阳性干血片标本患者SNaPshot技术和测序技术的结果完全吻合,有2例复合突变,3例纯合突变,显示SNaPshot技术高度的特异性和灵敏度。结论采用SNaPshot技术检测CYP21A2基因具有高通量、准确、对样本容受性高的特点,便于临床应用。Objective： To establish the method for rapid gene diagnosis of congenital adrenal hyperplasia （CAH） with 21-hydroxylase deficiency （21-OHD） . Methods： Nest amplification of Chinese population 21-hydroxylase deficiency the common mutation hot Exon3 Del 8bp, Q318X, R356W, the IVS2 A/C〉G, I172N, P30L and V281L of seven sites were performed using multiplex PCR technology. Afterwards, the sequence-specific probes interrogates each locus and labels it at the 3＇ end using fluorescent dideoxynucleotide chemistry by the SNaPshot Multiplex Kit, the resulting products were then analyzed and genotyped in the presence of a fifth-dye-labeled size standard. Results： A rapid detection method of CYP21A2 gene set to meet quality control standards was established, the results of Snapshot technology was consistent with the results of the sequencing technology in five positive dried blood specimens. Two cases were complex mutation, three cases were homozygous mutation. It is displayed a high degree of specificity and sensitivity of the Snapshot technology. Conclusion： The SNaPshot technology of CYP21A2 gene detection had a high-throughput, accurate, and sample characteristics of high receptivity for clinical application.

关 键 词：肾上腺皮质增生症 SNaPshot技术 CYP21A2基因 测序技术 

分 类 号：R586[医药卫生—内分泌]