Massively parallel sequencing, aCGH, and RNA-Seq technologies provide a comprehensive molecular diagnosis of Fanconi anemia
Massively parallel sequencing, aCGH, and RNA-Seq technologies provide a comprehensive molecular diagnosis of Fanconi anemia
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DOI：
10.1182/blood-2012-12-474585
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发表时间：
2013-05-30
期刊：
影响因子：
20.3
通讯作者：
Ostrander, Elaine A.
中科院分区：
文献类型：
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作者：
Chandrasekharappa, Settara C.;Lach, Francis P.;Ostrander, Elaine A.
目前检测范可尼贫血（FA）疑似患者突变的方法效率低下，经常遗漏突变。我们将DNA测序和基因组捕获的最新进展应用于FA的诊断。具体来说，我们使用定制的分子倒置探针或truseq富集寡核苷酸来捕获FA和相关基因，包括内含子，并对来自洛克菲勒大学国际范可尼贫血登记处的27个样本进行测序。DNA测序辅以定制阵列比较基因组杂交（aCGH）和RNA测序（RNA-seq）分析。aCGH鉴定了4个不同FA基因的缺失/重复。RNA-seq分析显示，缺乏等位基因特异性表达与错义、同义和内含子深层变异引起的缺失和剪接缺陷相关。truseq靶向捕获、aCGH和RNA-seq的结合使我们能够识别所有27个家族的互补组和双等位基因种系突变：FANCA(7)、FANCB(3)、FANCC(3)、FANCD1(1)、FANCD2(3)、FANCF(2)、FANCG(2)、FANCI(1)、FANCJ(2)和FANCL(3)。FANCC突变通常是阿什肯纳兹犹太人（AJ）血统患者发生FA的原因，我们在2名AJ血统患者中发现了2个新的FANCC突变。我们在这里描述了一种有效的FA分子诊断策略。
Current methods for detecting mutations in Fanconi anemia (FA)-suspected patients are inefficient and often miss mutations. We have applied recent advances in DNA sequencing and genomic capture to the diagnosis of FA. Specifically, we used custom molecular inversion probes or TruSeq-enrichment oligos to capture and sequence FA and related genes, including introns, from 27 samples from the International Fanconi Anemia Registry at The Rockefeller University. DNA sequencing was complemented with custom array comparative genomic hybridization (aCGH) and RNA sequencing (RNA-seq) analysis. aCGH identified deletions/duplications in 4 different FA genes. RNA-seq analysis revealed lack of allele specific expression associated with a deletion and splicing defects caused by missense, synonymous, and deep-in-intron variants. The combination of TruSeq-targeted capture, aCGH, and RNA-seq enabled us to identify the complementation group and biallelic germline mutations in all 27 families: FANCA (7), FANCB (3), FANCC (3), FANCD1 (1), FANCD2 (3), FANCF (2), FANCG (2), FANCI (1), FANCJ (2), and FANCL (3). FANCC mutations are often the cause of FA in patients of Ashkenazi Jewish (AJ) ancestry, and we identified 2 novel FANCC mutations in 2 patients of AJ ancestry. We describe here a strategy for efficient molecular diagnosis of FA.