Cargando…

A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax

BACKGROUND: Primary spontaneous pneumothorax (PSP) or pulmonary cysts is one of the manifestations of Birt-Hogg-Dube syndrome (BHDS) that is caused by heterozygous mutations in FLCN gene. Most of the mutations are SNVs and small indels, and there are also approximately 10 % large intragenic deletion...

Descripción completa

Detalles Bibliográficos
Autores principales: Zhang, Xinxin, Ma, Dehua, Zou, Wei, Ding, Yibing, Zhu, Chengchu, Min, Haiyan, Zhang, Bin, Wang, Wei, Chen, Baofu, Ye, Minhua, Cai, Minghui, Pan, Yanqing, Cao, Lei, Wan, Yueming, Jin, Yu, Gao, Qian, Yi, Long
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2016
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4882857/
https://www.ncbi.nlm.nih.gov/pubmed/27229674
http://dx.doi.org/10.1186/s12931-016-0377-9
_version_ 1782434188277841920
author Zhang, Xinxin
Ma, Dehua
Zou, Wei
Ding, Yibing
Zhu, Chengchu
Min, Haiyan
Zhang, Bin
Wang, Wei
Chen, Baofu
Ye, Minhua
Cai, Minghui
Pan, Yanqing
Cao, Lei
Wan, Yueming
Jin, Yu
Gao, Qian
Yi, Long
author_facet Zhang, Xinxin
Ma, Dehua
Zou, Wei
Ding, Yibing
Zhu, Chengchu
Min, Haiyan
Zhang, Bin
Wang, Wei
Chen, Baofu
Ye, Minhua
Cai, Minghui
Pan, Yanqing
Cao, Lei
Wan, Yueming
Jin, Yu
Gao, Qian
Yi, Long
author_sort Zhang, Xinxin
collection PubMed
description BACKGROUND: Primary spontaneous pneumothorax (PSP) or pulmonary cysts is one of the manifestations of Birt-Hogg-Dube syndrome (BHDS) that is caused by heterozygous mutations in FLCN gene. Most of the mutations are SNVs and small indels, and there are also approximately 10 % large intragenic deletions and duplications of the mutations. These molecular findings are generally obtained by disparate methods including Sanger sequencing and Multiple Ligation-dependent Probe Amplification in the clinical laboratory. In addition, as a genetically heterogeneous disorder, PSP may be caused by mutations in multiple genes include FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 genes. For differential diagnosis, these genes should also be screened which makes the diagnostic procedure more time-consuming and labor-intensive. METHODS: Forty PSP patients were divided into 2 groups. Nineteen patients with different pathogenic mutations of FLCN previously identified by conventional Sanger sequencing and MLPA were included in test group, 21 random PSP patients without any genetic screening were included in blinded sample group. 7 PSP genes including FLCN, FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 were designed and enriched by Haloplex system, sequenced on a Miseq platform and analyzed in the 40 patients to evaluate the performance of the targeted-NGS method. RESULTS: We demonstrated that the full spectrum of genes associated with pneumothorax including FLCN gene mutations can be identified simultaneously in multiplexed sequence data. Noteworthy, by our in-house copy number analysis of the sequence data, we could not only detect intragenic deletions, but also determine approximate deletion junctions simultaneously. CONCLUSIONS: NGS based Haloplex target enrichment technology is proved to be a rapid and cost-effective screening strategy for the comprehensive molecular diagnosis of BHDS in PSP patients, as it can replace Sanger sequencing and MLPA by simultaneously detecting exonic and intronic SNVs, small indels, large intragenic deletions and determining deletion junctions in PSP-related genes.
format Online
Article
Text
id pubmed-4882857
institution National Center for Biotechnology Information
language English
publishDate 2016
publisher BioMed Central
record_format MEDLINE/PubMed
spelling pubmed-48828572016-05-28 A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax Zhang, Xinxin Ma, Dehua Zou, Wei Ding, Yibing Zhu, Chengchu Min, Haiyan Zhang, Bin Wang, Wei Chen, Baofu Ye, Minhua Cai, Minghui Pan, Yanqing Cao, Lei Wan, Yueming Jin, Yu Gao, Qian Yi, Long Respir Res Research BACKGROUND: Primary spontaneous pneumothorax (PSP) or pulmonary cysts is one of the manifestations of Birt-Hogg-Dube syndrome (BHDS) that is caused by heterozygous mutations in FLCN gene. Most of the mutations are SNVs and small indels, and there are also approximately 10 % large intragenic deletions and duplications of the mutations. These molecular findings are generally obtained by disparate methods including Sanger sequencing and Multiple Ligation-dependent Probe Amplification in the clinical laboratory. In addition, as a genetically heterogeneous disorder, PSP may be caused by mutations in multiple genes include FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 genes. For differential diagnosis, these genes should also be screened which makes the diagnostic procedure more time-consuming and labor-intensive. METHODS: Forty PSP patients were divided into 2 groups. Nineteen patients with different pathogenic mutations of FLCN previously identified by conventional Sanger sequencing and MLPA were included in test group, 21 random PSP patients without any genetic screening were included in blinded sample group. 7 PSP genes including FLCN, FBN1, COL3A1, CBS, SERPINA1 and TSC1/TSC2 were designed and enriched by Haloplex system, sequenced on a Miseq platform and analyzed in the 40 patients to evaluate the performance of the targeted-NGS method. RESULTS: We demonstrated that the full spectrum of genes associated with pneumothorax including FLCN gene mutations can be identified simultaneously in multiplexed sequence data. Noteworthy, by our in-house copy number analysis of the sequence data, we could not only detect intragenic deletions, but also determine approximate deletion junctions simultaneously. CONCLUSIONS: NGS based Haloplex target enrichment technology is proved to be a rapid and cost-effective screening strategy for the comprehensive molecular diagnosis of BHDS in PSP patients, as it can replace Sanger sequencing and MLPA by simultaneously detecting exonic and intronic SNVs, small indels, large intragenic deletions and determining deletion junctions in PSP-related genes. BioMed Central 2016-05-27 2016 /pmc/articles/PMC4882857/ /pubmed/27229674 http://dx.doi.org/10.1186/s12931-016-0377-9 Text en © Zhang et al. 2016 Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0 International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, and reproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.
spellingShingle Research
Zhang, Xinxin
Ma, Dehua
Zou, Wei
Ding, Yibing
Zhu, Chengchu
Min, Haiyan
Zhang, Bin
Wang, Wei
Chen, Baofu
Ye, Minhua
Cai, Minghui
Pan, Yanqing
Cao, Lei
Wan, Yueming
Jin, Yu
Gao, Qian
Yi, Long
A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title_full A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title_fullStr A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title_full_unstemmed A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title_short A rapid NGS strategy for comprehensive molecular diagnosis of Birt-Hogg-Dubé syndrome in patients with primary spontaneous pneumothorax
title_sort rapid ngs strategy for comprehensive molecular diagnosis of birt-hogg-dubé syndrome in patients with primary spontaneous pneumothorax
topic Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4882857/
https://www.ncbi.nlm.nih.gov/pubmed/27229674
http://dx.doi.org/10.1186/s12931-016-0377-9
work_keys_str_mv AT zhangxinxin arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT madehua arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zouwei arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT dingyibing arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zhuchengchu arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT minhaiyan arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zhangbin arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT wangwei arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT chenbaofu arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT yeminhua arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT caiminghui arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT panyanqing arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT caolei arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT wanyueming arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT jinyu arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT gaoqian arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT yilong arapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zhangxinxin rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT madehua rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zouwei rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT dingyibing rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zhuchengchu rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT minhaiyan rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT zhangbin rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT wangwei rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT chenbaofu rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT yeminhua rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT caiminghui rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT panyanqing rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT caolei rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT wanyueming rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT jinyu rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT gaoqian rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax
AT yilong rapidngsstrategyforcomprehensivemoleculardiagnosisofbirthoggdubesyndromeinpatientswithprimaryspontaneouspneumothorax