An improved Diagnostic PCR Assay for identification of Cryptic Heterozygosity for CGG Triplet Repeat Alleles in the Fragile X Gene (FMR1).
An improved Diagnostic PCR Assay for identification of Cryptic Heterozygosity for CGG Triplet Repeat Alleles in the Fragile X Gene (FMR1).
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DOI：
10.1186/1755-8166-1-5
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发表时间：
2008-04-08
影响因子：
1.3
通讯作者：
Slater, Howard R.
中科院分区：
文献类型：
--
作者：
Khaniani, Mahmoud S.;Kalitsis, Paul;Burgess, Trent;Slater, Howard R.
脆性X染色体综合征（OMIM #300624）是最常见的、公认的、可遗传的导致智力迟钝的原因。由于该病的发病频率相对较高，早期发现的好处，以及后代遗传扩大致病性突变风险为1 / 2的相关携带者的识别，有必要进行广泛的检测。然而，由于缺乏一种单一、简单的实验室测试，对智力迟钝者进行具有成本效益的筛查一直受到阻碍。目前，通过简单的高通量PCR检测，可以在男性和大多数女性中排除脆性X综合征。由于PCR检测的灵敏度有限，我们发现在我们的诊断服务中，大约40%的女性表现为纯合子，需要进行劳动密集型和昂贵的Southern blot检测来区分这些女性携带一个正常等位基因和一个扩展等位基因。我们描述了一个改进的PCR测试，它显示了一个高水平的精度，允许等位基因不同的一个单一的三联体被解决。使用新的检测方法，我们检测到46/83（53%）以前标记为纯合子的隐杂合子。该分析还扩大了重复序列的扩增范围，在男性中扩增到170个CGG重复序列，在女性中扩增到130个CGG重复序列。结合高精度，该方法还提高了正常等位基因（CGG重复数< 45）与灰色区域等位基因（45 < CGG重复数< 54）和灰色区域等位基因与小前突变（55 < CGG重复数< 100）的区分。使用这种PCR测试提供了显著提高的精度和长等位基因的扩增。所需的后续南印迹试验次数减少（最多50%），从而改善了周转时间和成本。
Fragile X syndrome (OMIM #300624) is the most common, recognised, heritable cause of mental retardation. Widespread testing is warranted by the relatively high frequency of the disorder, the benefits of early detection and the identification of related carriers whose offspring are at a 1 in 2 risk of inheriting the expanded pathogenic mutation. However, cost-effective screening of mentally retarded individuals has been impeded by the lack of a single, simple laboratory test. Currently, Fragile X syndrome can be excluded in males and a majority of females using a simple high-throughput PCR test. Due to the limited sensitivity of the PCR test, we find in our diagnostic service that approximately 40% of females appear homozygous and a labour intensive and expensive Southern blot test is required to distinguish these from females carrying one normal allele and an expanded allele. We describe an improved PCR test which displays a high level of precision allowing alleles differing by a single triplet to be resolved. Using the new assay, we detected 46/83 (53%) cryptic heterozygotes previously labelled as homozygotes. The assay also extended the range of repeats amplifiable, up to 170 CGG repeats in males and 130 CGG repeats in females. Combined with the high precision, the assay also improves discrimination of normal (CGG repeats < 45) from grey zone (45 < CGG repeats < 54) alleles and grey zone alleles from small premutations (55 < CGG repeats < 100). Use of this PCR test provides significantly improved precision and amplification of longer alleles. The number of follow-up Southern blot tests required is reduced (up to 50%) with consequent improvement in turnaround time and cost.
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影响因子：
64.5
作者：
FU, YH;KUHL, DPA;CASKEY, CT
通讯作者：
CASKEY, CT
影响因子：
6.8
作者：
Houdayer, C;Lemonnier, A;Feldmann, D
通讯作者：
Feldmann, D
影响因子：
4.8
作者：
KANG, SM;OHSHIMA, K;WELLS, RD
通讯作者：
WELLS, RD
影响因子：
8.8
作者：
Strom, Charles M.;Crossley, Beryl;Sun, Weimin
通讯作者：
Sun, Weimin
影响因子：
6.8
作者：
Hecimovic, S;Vlasic, J;Pavelic, K
通讯作者：
Pavelic, K