Comprehensive assessment of sequence variation within the copy number variable defensin cluster on 8p23 by target enriched in-depth 454 sequencing

In highly copy number variable (CNV) regions such as the human defensin gene locus, comprehensive assessment of sequence variations is challenging. PCR approaches are practically restricted to tiny fractions, and next-generation sequencing (NGS) approaches of whole individual genomes e.g. by the 100...

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Bibliographic Details
Published in:BMC genomics 2011-05, Vol.12 (1), p.243-243, Article 243
Main Authors: Taudien, Stefan, Szafranski, Karol, Felder, Marius, Groth, Marco, Huse, Klaus, Raffaelli, Francesca, Petzold, Andreas, Zhang, Xinmin, Rosenstiel, Philip, Hampe, Jochen, Schreiber, Stefan, Platzer, Matthias
Format: Article
Language:English
Subjects:
Base Sequence
Cell Line
Chromosomes, Human, Pair 8 - genetics
Defensins - genetics
DNA Copy Number Variations
Gene Dosage
Gene Frequency
Genetic Variation
Genomes
Haplotypes
High-Throughput Nucleotide Sequencing
Humans
Male
Multigene Family
Nucleotide sequence
Physiological aspects
Polymerase chain reaction
Sequence Analysis, DNA
Single nucleotide polymorphisms
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Summary:In highly copy number variable (CNV) regions such as the human defensin gene locus, comprehensive assessment of sequence variations is challenging. PCR approaches are practically restricted to tiny fractions, and next-generation sequencing (NGS) approaches of whole individual genomes e.g. by the 1000 Genomes Project is confined by an affordable sequence depth. Combining target enrichment with NGS may represent a feasible approach. As a proof of principle, we enriched a ~850 kb section comprising the CNV defensin gene cluster DEFB, the invariable DEFA part and 11 control regions from two genomes by sequence capture and sequenced it by 454 technology. 6,651 differences to the human reference genome were found. Comparison to HapMap genotypes revealed sensitivities and specificities in the range of 94% to 99% for the identification of variations.Using error probabilities for rigorous filtering revealed 2,886 unique single nucleotide variations (SNVs) including 358 putative novel ones. DEFB CN determinations by haplotype ratios were in agreement with alternative methods. Although currently labor extensive and having high costs, target enriched NGS provides a powerful tool for the comprehensive assessment of SNVs in highly polymorphic CNV regions of individual genomes. Furthermore, it reveals considerable amounts of putative novel variations and simultaneously allows CN estimation.
ISSN:1471-2164
1471-2164
DOI:10.1186/1471-2164-12-243