【摘要】 目的建立变性高效液相色谱筛查G6PD基因变异的技术。观察广东汉族和壮族G6PD基因变异型的分布。方法应用变性高效液相色谱技术筛查广东汉族和壮族G6PD基因变异型。与ARMS及DNA测序比较,考察其准确性、灵敏度、效率及耗资。结果在73例G6PD缺陷者中发现G6PD1388G→A20例、1376G→T24例、95A→G6例、1311C→T8例、1024C→T3例、392G→T2例、871G→A1例、1376G→A/95A→G4例、1388G→A/95A→G3例、1311C→T/95A→G2例,与DNA测序结果吻合;在酶学正常对照组中发现男1例女2例1388突变。结论变性高效液相色谱技术对G6PD基因变异型的筛查具有准确、高效、半自动化及经济等优点。尤其对无症状女性杂合子及临床Ⅳ型个体的筛查具有重要意义。更多还原

【Abstract】 Objective Of denaturing high performance liquid chromatography (DHPLC), a technique platform was developed for screening G6PD deficient variants. Methods When applied to screen and identify the G6PD deficient variants from 124 patients who come from 11 nations in China,the DHPLC was compared with amplification refractory mutation system (ARMS) or DNA sequence technique;and assessed carefully in its accuracy, sensitivity, efficiency and the cost of experiment. Results The G6PD-deficient variants, such as 1388 G→A(36/124 cases), 1376 G→T(35),95 A→ G(14), 1024 C→T (3),392 G→T(4), 871 G→A/1311 C→ T/IVS XI +93 t→c(9) , 871 G→A (1), 1311 C→ T/IVS XI +93 t→c (4),1376 G→T/1388 G→A (1) and so on, were characterized as sharp peaks by DHPLC and verified by DNA sequence. Further, the standard chromatograms were put into database for 8 kinds of common G6PD deficient variants in Chinese populations. And also DHPLC found 3 G6PD variants (1388 G→A) from 103 negative controls. With taking 8.8 minutes and costing 1 dollar for each sample,DHPLC successfully detected and identified 34 heterozygous females from patients with G6PD deficiency. However, ARMS checked 83 positive controls but got 12 false G6PD mutants, of which 5 were false positive, 7 false negative. Above results show that DHPLC sounds like to be more convenience, sensitive and accurate than ARMS and DNA sequence techniques for checking G6PD mutants. Conclusion DHPLC is of great advantage to screen the G6PD deficient variants with accuracy, convenience, automation and less cost,and significantly to identify the female heterozygote and clinical type Ⅳ individuals with G6PD deficiency.更多还原