一种专门设计的多基因检测板有助于肝内胆汁淤积症患儿的基因诊断：已知大片段插入/缺失的同步检测
A Specially Designed Multi-Gene Panel Facilitates Genetic Diagnosis in Children with Intrahepatic Cholestasis: Simultaneous Test of Known Large Insertions/Deletions.
作者信息
Wang Neng-Li, Lu Yu-Lan, Zhang Ping, Zhang Mei-Hong, Gong Jing-Yu, Lu Yi, Xie Xin-Bao, Qiu Yi-Ling, Yan Yan-Yan, Wu Bing-Bing, Wang Jian-She
机构信息
Department of Pediatrics, Jinshan Hospital of Fudan University, Shanghai, China.
The Molecular Genetic Diagnosis Center, Shanghai Key Lab of Birth Defects, Pediatrics Research Institute, Children's Hospital of Fudan University, Shanghai, China.
出版信息
PLoS One. 2016 Oct 5;11(10):e0164058. doi: 10.1371/journal.pone.0164058. eCollection 2016.
BACKGROUND AND AIMS
Large indels are commonly identified in patients but are not detectable by routine Sanger sequencing and panel sequencing. We specially designed a multi-gene panel that could simultaneously test known large indels in addition to ordinary variants, and reported the diagnostic yield in patients with intrahepatic cholestasis.
METHODS
The panel contains 61 genes associated with cholestasis and 25 known recurrent large indels. The amplicon library was sequenced on Ion PGM system. Sequencing data were analyzed using a routine data analysis protocol and an internal program encoded for large indels test simultaneously. The validation phase was performed using 54 patients with known genetic diagnosis, including 5 with large insertions. At implement phase, 141 patients with intrahepatic cholestasis were evaluated.
RESULTS
At validation phase, 99.6% of the variations identified by Sanger sequencing could be detected by panel sequencing. Following the routine protocol, 99.8% of false positives could be filtered and 98.8% of retained variations were true positives. Large insertions in the 5 patients with known genetic diagnosis could be correctly detected using the internal program. At implementation phase, 96.9% of the retained variations, following the routine protocol, were confirmed to be true. Twenty-nine patients received a potential genetic diagnosis when panel sequencing data were analyzed using the routine protocol. Two additional patients, who were found to harbor large insertions in SLC25A13, obtained a potential genetic diagnosis when sequencing data were further analyzed using the internal program. A total of 31 (22.0%) patients obtained a potential genetic diagnosis. Nine different genetic disorders were diagnosed, and citrin deficiency was the commonest.
CONCLUSION
Specially designed multi-gene panel can correctly detect large indels simultaneously. By using it, we assigned a potential genetic diagnosis to 22.0% of patients with intrahepatic cholestasis.
背景与目的
大片段插入缺失在患者中较为常见，但常规桑格测序和基因panel测序无法检测到。我们专门设计了一个多基因panel，除了能检测普通变异外，还能同时检测已知的大片段插入缺失，并报告了其在肝内胆汁淤积症患者中的诊断率。
方法
该panel包含61个与胆汁淤积相关的基因以及25个已知的复发性大片段插入缺失。扩增子文库在Ion PGM系统上进行测序。测序数据使用常规数据分析方案和为大片段插入缺失检测同时编码的内部程序进行分析。验证阶段使用54例已知基因诊断的患者进行，其中包括5例有大片段插入的患者。在实施阶段，对141例肝内胆汁淤积症患者进行了评估。
结果
在验证阶段，桑格测序鉴定出的变异中有99.6%可通过panel测序检测到。按照常规方案，99.8%的假阳性可被过滤，98.8%保留的变异为真阳性。使用内部程序可正确检测出5例已知基因诊断患者中的大片段插入。在实施阶段，按照常规方案，96.9%保留的变异被确认为真实变异。使用常规方案分析panel测序数据时，29例患者获得了潜在的基因诊断。另外2例在SLC25A13中发现有大片段插入的患者，在使用内部程序进一步分析测序数据时获得了潜在的基因诊断。共有31例（22.0%）患者获得了潜在的基因诊断。诊断出9种不同的遗传疾病，其中柠檬酸转运蛋白缺乏症最为常见。
结论
专门设计的多基因panel能够同时正确检测大片段插入缺失。通过使用该panel，我们为22.0%的肝内胆汁淤积症患者做出了潜在的基因诊断。
Zotero 插件