验证纳米孔长读数测序技术,以确定 X 连锁视网膜色素变性患者的 RPGR ORF15 基因型。

IF 3.7 2区 生物学 Q2 BIOCHEMISTRY & MOLECULAR BIOLOGY
Christel Vaché, Valérie Faugère, David Baux, Luke Mansard, Charles Van Goethem, Claire-Marie Dhaenens, Olivier Grunewald, Isabelle Audo, Christina Zeitz, Isabelle Meunier, Béatrice Bocquet, Mireille Cossée, Anne Bergougnoux, Vasiliki Kalatzis, Anne-Françoise Roux
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引用次数: 0

摘要

X 连锁色素性视网膜炎(XLRP)的特征是进行性视力丧失,男性患者会导致法定失明,而女性携带者的严重程度则很宽泛。视网膜色素变性 GTPase 调节基因(RPGR)的致病性改变是导致 70% 以上 XLRP 病例的原因。在视网膜中,RPGRORF15 转录本包括一个称为 ORF15 的末端外显子,在大多数 RPGR-XLRP 病例中,该外显子都发生了改变。遗憾的是,由于 ORF15 的序列重复性很高,因此对于分子诊断实验室来说,ORF15 的测序是一个相当大的挑战。不过,在最近的一项初步研究中,Yahya 等人报告了一种长读数测序方法,似乎很有希望。本研究的目的是验证这种新的测序策略,并将其整合到常规筛查工作流程中。为此,我们对携带 32 种不同致病性 ORF15 变异(包括位于外显子高度重复区域的 20 种)的男性和女性个体的 52 份基因组 DNA 样本进行了掩蔽测试。经过生物信息学分析,我们发现后者在男性中的检出率为 80-85%,在女性中的检出率为 60-80%。在对 ORF15 长读数进行补充性目视检查后,这两种状态的检出率均提高到了 100%。根据这些结果,并考虑到ORF15在XLRP中的致病变异频率,我们建议在对诊断符合XLRP的受试者进行任何其他测序方法之前,应系统地考虑对ORF15进行长读数筛查。
本文章由计算机程序翻译,如有差异,请以英文原文为准。

Validation of Nanopore long-read sequencing to resolve RPGR ORF15 genotypes in individuals with X-linked retinitis pigmentosa.

Validation of Nanopore long-read sequencing to resolve RPGR ORF15 genotypes in individuals with X-linked retinitis pigmentosa.

X-linked retinitis pigmentosa (XLRP) is characterized by progressive vision loss leading to legal blindness in males and a broad severity spectrum in carrier females. Pathogenic alterations of the retinitis pigmentosa GTPase regulator gene (RPGR) are responsible for over 70% of XLRP cases. In the retina, the RPGRORF15 transcript includes a terminal exon, called ORF15, that is altered in the large majority of RPGR-XLRP cases. Unfortunately, due to its highly repetitive sequence, ORF15 represents a considerable challenge in terms of sequencing for molecular diagnostic laboratories. However, in a recent preliminary work Yahya et al. reported a long-read sequencing approach seeming promising. Here, the aim of the study was to validate and integrate this new sequencing strategy in a routine screening workflow. For that purpose, we performed a masked test on 52 genomic DNA samples from male and female individuals carrying 32 different pathogenic ORF15 variations including 20 located in the highly repetitive region of the exon. For the latter, we have obtained a detection rate of 80-85% in males and 60-80% in females after bioinformatic analyses. These numbers raised to 100% for both status after adding a complementary visual inspection of ORF15 long-reads. In accordance with these results, and considering the frequency of ORF15 pathogenic variations in XLRP, we suggest that a long-read screening of ORF15 should be systematically considered before any other sequencing approach in subjects with a diagnosis compatible with XLRP.

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来源期刊
European Journal of Human Genetics
European Journal of Human Genetics 生物-生化与分子生物学
CiteScore
9.90
自引率
5.80%
发文量
216
审稿时长
2 months
期刊介绍: The European Journal of Human Genetics is the official journal of the European Society of Human Genetics, publishing high-quality, original research papers, short reports and reviews in the rapidly expanding field of human genetics and genomics. It covers molecular, clinical and cytogenetics, interfacing between advanced biomedical research and the clinician, and bridging the great diversity of facilities, resources and viewpoints in the genetics community. Key areas include: -Monogenic and multifactorial disorders -Development and malformation -Hereditary cancer -Medical Genomics -Gene mapping and functional studies -Genotype-phenotype correlations -Genetic variation and genome diversity -Statistical and computational genetics -Bioinformatics -Advances in diagnostics -Therapy and prevention -Animal models -Genetic services -Community genetics
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