应用PCR-反向点杂交法检测β-地中海贫血基因点突变  

Detection of point mutations of β-thalassemia gene by PCR-reverse dot blot

作　　者：吕燕波[1] 潘兴华[1] 徐庆玲[2] 刘林[3] 邓永丽[1] 王金祥[1] 石琳琳[1] 

机构地区：[1]成都军区昆明总医院临床实验科,昆明650032 [2]成都军区昆明总医院儿科,昆明650032 [3]成都军区昆明总医院血液内科,昆明650032

出　　处：《西南国防医药》2010年第7期717-719,共3页Medical Journal of National Defending Forces in Southwest China

基　　金：云南省面上项目(2009ZC150M);成都军区面上项目(MB09028)

摘　　要：目的对疑似β-地中海贫血的4例患者进行基因诊断。方法针对β-珠蛋白基因上17个位点突变,使用特异性带有生物素标记的引物进行PCR扩增,扩增产物与固定在膜条上的寡核苷酸探针进行杂交,并通过一系列显色反应,明确检测样品在17个位点是否发生了突变,以及是否为突变纯合子或杂合子。结果 1号和2号患者均为CD17M(A→T)/N,βEM(GAG→AAG)/N双重杂合子;D3号患者为IVS-Ⅱ-654M(C→T)/N杂合子;D4号患者为IVS-Ⅱ-654M/IVS-Ⅱ-654M(C→T)纯合子。结论利用PCR-反向点杂交法技术,能对β-地中海贫血患者进行基因诊断,对其产前诊断、临床治疗以及携带者检出具有重要意义。Objective To perform gene diagnosis on four subjects with suspected β - thalassemia. Methods Biotin labeled primers specific to seventeen mutations of β - globulin gene were used to amplify its DNA fragments. A set of oligonucleotide probes were immobilized on the nylon membrane to capture biotin labeled amplification products by hybridization. Then series of color reactions were conducted to determine whether seventeen - gene locus of the samples mutated and whether the mutation gene was homozygote or heterozygote. Results Patient No. 1 and No. 2 were all the dual heterozygote of CD17M （ A→T）/N, βEM （ GAG→AAG）/N. Patient D3 was a heterozygote of IVS - Ⅱ - 654M （ C→T）/N. Patient D4 was a homozygote of IVS - Ⅱ - 654M/IVS - Ⅱ - 654M （C→T）. Conclusion PCR - reverse dot blot is a convenient, high performance and reliable method for the genetic diagnosis of β - thalassemia, and it is useful for prenatal diagnosis, clinical treatment and carrier screening of this disease.

关 键 词：Β-地中海贫血 点突变 PCR-反向点杂交 

分 类 号：R556.61[医药卫生—血液循环系统疾病]