Hüebner Claudia, Petermann Ivonne, Browning Brian L, Shelling Andrew N, Ferguson Lynnette R
Discipline of Nutrition, Faculty of Medical and Health Sciences, University of Auckland, Auckland, New Zealand.
Cancer Epidemiol Biomarkers Prev. 2007 Jun;16(6):1185-92. doi: 10.1158/1055-9965.EPI-06-0759.
Accurate measurement of allele frequencies between population groups with differing sensitivities to disease is fundamental to genetic epidemiology. Genotyping errors can markedly influence the biological conclusions of a study. This issue may be especially important now there is increasing recognition of triallelic single nucleotide polymorphisms (SNPs) in the genome and their possible role in diseases like inflammatory bowel disease. For example, the MDR1 (ABCB1) SNP G2677/T/A was, like many other triallelic SNPs, originally described as diallelic. Here, we report a comprehensive analyses of estimated allele frequencies of this SNP in a set of 73 human DNA samples, comparing six commonly used genotyping methods (Applied Biosystems Taqman, Roche LightCycler melting analysis, allelic discrimination PCR, DNA sequencing, Sequenom, and RFLP) from the angle of their error potential. Only Sequenom and DNA sequencing provided accurate measurements, if we had not had prior knowledge of the triallelic nature of this SNP. The other tested methods (with the exception of LightCycler) failed to show any indication of the presence of the rare third A- allele in a diallelic assay. Although most of the errors were due to the inability to detect the third allele, all methods except Sequenom and sequencing produced errors for the detection of the two common alleles G and T (LightCycler, 6 errors; PCR, 4 errors; RFLP, 2 errors; Taqman, 1 error). There is considerable variability in the reported frequencies of the different alleles of the MDR1 G2677/T/A SNP, and the role of this SNP in the etiology of inflammatory bowel disease has been controversial. Our data emphasize the importance of choosing the appropriate method for SNP detection and lead us to suggest that part of the previously reported variation may reflect artifacts associated with the different genotyping methodologies used. The failure to recognize the triallic nature of a SNP may lead to underestimations of real genetic associations.
准确测量对疾病敏感度不同的人群组之间的等位基因频率是遗传流行病学的基础。基因分型错误会显著影响研究的生物学结论。鉴于目前人们越来越认识到基因组中的三等位基因单核苷酸多态性（SNP）及其在炎症性肠病等疾病中的可能作用，这个问题可能尤为重要。例如，MDR1（ABCB1）SNP G2677/T/A与许多其他三等位基因SNP一样，最初被描述为双等位基因。在此，我们报告了对一组73个人类DNA样本中该SNP估计等位基因频率的全面分析，从其潜在误差的角度比较了六种常用的基因分型方法（应用生物系统公司的Taqman、罗氏LightCycler熔解分析、等位基因鉴别PCR、DNA测序、Sequenom和RFLP）。如果我们事先不知道该SNP的三等位基因性质，只有Sequenom和DNA测序能提供准确测量。其他测试方法（LightCycler除外）在双等位基因检测中未能显示出存在罕见的第三个A等位基因的任何迹象。尽管大多数错误是由于无法检测到第三个等位基因，但除Sequenom和测序外的所有方法在检测两个常见等位基因G和T时也产生了错误（LightCycler，6个错误；PCR，4个错误；RFLP，2个错误；Taqman，1个错误）。MDR1 G2677/T/A SNP不同等位基因的报告频率存在相当大的差异，并且该SNP在炎症性肠病病因学中的作用一直存在争议。我们的数据强调了选择合适的SNP检测方法的重要性，并使我们认为先前报告的部分变异可能反映了与所使用的不同基因分型方法相关的假象。未能认识到SNP的三等位基因性质可能导致对真实遗传关联的低估。
