Rapid identification of female carriers of DMD/BMD by quantitative real-time PCR

被引:44
作者
Joncourt, F [1 ]
Neuhaus, B [1 ]
Jostarndt-Foegen, K [1 ]
Kleinle, S [1 ]
Steiner, B [1 ]
Gallati, S [1 ]
机构
[1] Childrens Univ Hosp, Div Human Genet, Inselspital, CH-3010 Bern, Switzerland
关键词
DMD; BMD; quantitative real-time PCR; light cycler; SYBR green I;
D O I
10.1002/humu.20007
中图分类号
Q3 [遗传学];
学科分类号
071007 ; 090102 ;
摘要
Recently developed PCR systems offer online-monitoring of amplification and allow simple and reliable DNA quantification. We have used the LightCycler system to develop a simple and rapid method for direct identification of female carriers of deletions and duplications in the dystrophin gene. The challenge resides in the ability to identify the presence of a deleted or duplicated allele over the background contributed by the normal allele. Quantification is based on the determination of the ratio between potentially deleted/duplicated dystrophin exons and non-deleted/-duplicated reference exons using the unspecific dsDNA-dye SYBRgreen I. In a retrospective study, we evaluated our method in female relatives of DMD/BMD patients with known carrier status by comparative analysis of deleted or duplicated versus non-deleted/-duplicated exons. Carrier status was accurately attributed in 100% of cases, the mean ratios being 0.52 +/- 0.12 for deletion carriers (expected value: 0.5) and 1.56 +/- 0.18 for duplication carriers (expected value: 1.5) vs. 1.022 +/- 0.17 for non-carriers (expected value: 1.0). The method proved to be simple, rapid, reliable, and cost-effective. It may be used for direct determination of deletions/duplications in potential DMD/BMD carriers and may easily be adapted for other genetic conditions involving deletions and duplications. (C) 2004 Wiley-Liss, Inc.
引用
收藏
页码:385 / 391
页数:7
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