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目的应用高分辨熔解曲线(high-resolution melting,HRM)分析技术检测中国南方常见的5种β地中海贫血基因突变,并初步探讨其临床应用价值。方法应用HRM法检测已知基因型的5种β地中海贫血突变(-28、CD17、CD41-42、CD26及IVS-2-654),评价该技术的准确性;然后对100例未知样本进行HRM分析,通过DNA测序进行验证,评价该技术的敏感性及特异性。结果 HRM法检测已知基因型的5种β地中海贫血突变结果与基因测序结果一致,诊断符合率100%。100例未知样本中,共检出5种常见β地贫杂合突变21例,包括3例-28、5例CD17、7例CD41-42、3例CD26及3例IVS-2-654突变,未发现相关位点的纯合型突变和双重杂合突变,上述结果与DNA测序结果完全相符。结论 HRM技术是一种简便快捷、准确敏感的基因检测技术,能准确检测广东省5种常见β地贫基因突变,有望成为β地贫大规模人群筛查新的实验方法。
Objective To detect the five common mutations of β-thalassemia in southern China by high-resolution melting (HRM) analysis and to explore its clinical value. Methods Five kinds of β-thalassemia mutations (-28, CD17, CD41-42, CD26 and IVS-2-654) of known genotypes were detected by HRM to evaluate the accuracy of the technique. Then, 100 unknown samples were subjected to HRM Analysis, validation by DNA sequencing to assess the sensitivity and specificity of the technology. Results Five mutations of β-thalassemia with known genotypes by HRM were consistent with the results of gene sequencing. The diagnostic coincidence rate was 100%. Among the 100 unknown samples, there were 21 cases of 5 common thalassemia heterozygous mutations including 3 cases -28,5 cases of CD17, 7 cases of CD41-42, 3 cases of CD26 and 3 cases of IVS-2-654 mutation, No homozygous mutation or double heterozygous mutation was found in the related sites. The above results are in good agreement with the results of DNA sequencing. Conclusion HRM is a simple and rapid, accurate and sensitive gene detection technique. It can accurately detect five common mutations of β-thalassemia gene in Guangdong province and is expected to become a new experimental method for large-scale β-thalassemia population screening.