Title
Simultaneous mutation and copy number variation (CNV) detection by multiplex PCR-based GS-FLX sequencing Simultaneous mutation and copy number variation (CNV) detection by multiplex PCR-based GS-FLX sequencing
Author
Faculty/Department
Faculty of Pharmaceutical, Biomedical and Veterinary Sciences . Biomedical Sciences
Publication type
article
Publication
New York, N.Y. ,
Subject
Human medicine
Source (journal)
Human mutation. - New York, N.Y.
Volume/pages
30(2009) :3 , p. 472-476
ISSN
1059-7794
ISI
000264011700025
Carrier
E
Target language
English (eng)
Full text (Publishers DOI)
Affiliation
University of Antwerp
Abstract
We evaluated multiplex PCR amplification as a front-end for high-throughput sequencing, to widen the applicability of massive parallel sequencers for the detailed analysis of complex genomes. Using multiplex PCR reactions, we sequenced the complete coding regions of seven genes implicated in peripheral neuropathies in 40 individuals on a GS-FLX genome sequencer (Roche). The resulting dataset showed highly specific and uniform amplification. Comparison of the GS-FLX sequencing data with the dataset generated by Sanger sequencing confirmed the detection of all variants present and proved the sensitivity of the method for mutation detection. In addition, we showed that we could exploit the multiplexed PCR amplicons to determine individual copy number variation (CNV), increasing the spectrum of detected variations to both genetic and genomic variants. We conclude that our straightforward procedure substantially expands the applicability of the massive parallel sequencers for sequencing projects of a moderate number of amplicons (50-500) with typical applications in resequencing exons in positional or functional candidate regions and molecular genetic diagnostics.
E-info
https://repository.uantwerpen.be/docman/iruaauth/e096ec/8566368809e.pdf
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