Tag Carmen G, Oberkanins Christian, Kriegshäuser Gernot, Ingram Catherine J E, Swallow Dallas M, Gressner Axel M, Ledochowski Maximilian, Weiskirchen Ralf
Institute of Clinical Chemistry and Pathobiochemistry, RWTH-University Hospital, D-52074 Aachen, Germany.
Clin Chim Acta. 2008 Jun;392(1-2):58-62. doi: 10.1016/j.cca.2008.03.006. Epub 2008 Mar 18.
Adult-type hypolactasia is a genetically determined inability to digest lactose after weaning. Two single-nucleotide polymorphisms (C-13910T, G-22018A) located upstream of the lactase gene (LCT) within the gene MCM6 are associated with the lactase persistence/non-persistence trait in patients of European descent. Therefore, the genotyping of these SNPs has been established as a diagnostic tool for adult-type hypolactasia. We have recently shown that several novel allelic variants located in close proximity to the C-13910T SNP interfere with the diagnostic accuracy of real-time PCR-based genotyping methods.
We describe here the validation of a comprehensive reverse-hybridization teststrip-based assay for the detection of common and novel LCT SNPs (C-13907G, C-13910T, T-13913C, G-13914A, T-13915G, and G-22018A). This assay is based on multiplex DNA amplification and ready-to-use membrane teststrips containing variant-specific oligonucleotide probes immobilized as an array of parallel lines.
We evaluated the novel reverse-hybridization StripAssay on 125 DNA samples in comparison to LightCycler analysis and sequencing. The outcome of StripAssay genotyping was found to be completely concordant with that obtained by sequencing.
The StripAssay represents an accurate and robust screening tool to identify multiple LCT/MCM6 variants in a rapid manner. It overcomes diagnostic pitfalls that were reported and allows the simultaneous genotyping of closely spaced LCT variant sites in a single-step diagnostic approach.
成人型乳糖不耐受是一种断奶后因基因决定而无法消化乳糖的情况。位于MCM6基因内乳糖酶基因（LCT）上游的两个单核苷酸多态性（C-13910T、G-22018A）与欧洲血统患者的乳糖酶持续性/非持续性特征相关。因此，这些单核苷酸多态性的基因分型已被确立为成人型乳糖不耐受的诊断工具。我们最近发现，位于C-13910T单核苷酸多态性附近的几个新的等位基因变体干扰了基于实时PCR的基因分型方法的诊断准确性。
我们在此描述一种基于反向杂交检测条的综合检测方法的验证，用于检测常见和新的LCT单核苷酸多态性（C-13907G、C-13910T、T-13913C、G-13914A、T-13915G和G-22018A）。该检测方法基于多重DNA扩增和即用型膜检测条，其中包含固定为平行线阵列的变体特异性寡核苷酸探针。
我们将新型反向杂交检测条法与LightCycler分析和测序相比，对125个DNA样本进行了评估。发现检测条法基因分型的结果与测序结果完全一致。
检测条法是一种准确且稳健的筛查工具，能够快速识别多个LCT/MCM6变体。它克服了已报道的诊断缺陷，并允许在单步诊断方法中对紧密间隔的LCT变体位点进行同时基因分型。
