Department of Laboratory Medicine, Graduate School, Kyung Hee University, Seoul, Korea.
Department of Laboratory Medicine, Seoul National University Hospital, Seoul National University College of Medicine, Seoul, Korea.
Ann Lab Med. 2021 Jul 1;41(4):394-400. doi: 10.3343/alm.2021.41.4.394.
Conventional diagnosis of fragile X syndrome (FXS) is based on a combination of fragment analysis (FA) and Southern blotting (SB); however, this diagnostic approach is time- and labor-intensive and has pitfalls such as the possibility of missing large number alleles. Triplet repeat primed PCR (TP-PCR) is a current alternative used to overcome these limitations. We evaluated the diagnostic usefulness of TP-PCR compared with the conventional diagnostic protocol consisting of FA and/or SB in terms of allele categorization, repeat number correlation, and zygosity concordance in female genetic carriers.
From November 2013 to March 2018, 458 patients (326 males, 132 females) were simultaneously examined using FA and/or SB and TP-PCR by detecting CGG repeat numbers in gene and diagnosed as per American College of Medical Genetics guidelines.
The TP-PCR results showed high concordance with the FA and/or SB results for all three aspects (allele categorization, repeat number correlation, and zygosity concordance in female genetic carriers). TP-PCR detected CGG expansions ≥200 in all full mutation (FM) allele cases in male patients, as well as both the normal allele (NL) and FM allele in female carriers. In premutation (PM) allele carriers, the TP-PCR results were consistent with the FA and/or SB results. In terms of zygosity concordance in female genetic carriers, 12 NL cases detected by TP-PCR showed a merged peak consisting of two close heterozygous peaks; however, this issue was resolved using a 10-fold dilution.
TP-PCR may serve as a reliable alternative method for FXS diagnosis.
脆性 X 综合征（FXS）的传统诊断基于片段分析（FA）和 Southern 印迹（SB）的组合；然而，这种诊断方法既耗时又费力，并且存在一些缺陷，例如可能会错过大量等位基因。三重复引物 PCR（TP-PCR）是一种当前的替代方法，可用于克服这些限制。我们评估了 TP-PCR 在等位基因分类、重复数相关性和女性遗传携带者的同型性一致性方面与包括 FA 和/或 SB 的传统诊断方案相比的诊断有用性。
从 2013 年 11 月到 2018 年 3 月，458 名患者（326 名男性，132 名女性）同时使用 FA 和/或 SB 和 TP-PCR 进行检查，通过检测基因中的 CGG 重复数，并根据美国医学遗传学学院的指南进行诊断。
TP-PCR 结果在所有三个方面（等位基因分类、重复数相关性和女性遗传携带者的同型性一致性）均与 FA 和/或 SB 结果高度一致。TP-PCR 在男性患者的所有完全突变（FM）等位基因病例中以及女性携带者的正常等位基因（NL）和 FM 等位基因中均检测到 CGG 扩展≥200。在前突变（PM）等位基因携带者中，TP-PCR 结果与 FA 和/或 SB 结果一致。在女性遗传携带者的同型性一致性方面，12 个通过 TP-PCR 检测到的 NL 病例显示出由两个接近的杂合峰组成的合并峰；然而，通过 10 倍稀释解决了这个问题。
TP-PCR 可能是 FXS 诊断的可靠替代方法。
