Ion Torrent sequencing for conducting genome-wide scans for mutation mapping analysis

Damerla, Rama Rao, Chatterjee, Bishwanath, Li, You, Francis, Richard J.B., Fatakia, Sarosh N., and Lo, Cecilia W. (2014) Ion Torrent sequencing for conducting genome-wide scans for mutation mapping analysis. Mammalian Genome, 25 (3-4). pp. 120-128.

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Mutation mapping in mice can be readily accomplished by genome wide segregation analysis of polymorphic DNA markers. In this study, we showed the efficacy of Ion Torrent next generation sequencing for conducting genome-wide scans to map and identify a mutation causing congenital heart disease in a mouse mutant, Bishu, recovered from a mouse mutagenesis screen. The Bishu mutant line generated in a C57BL/6J (B6) background was intercrossed with another inbred strain, C57BL/10J (B10), and the resulting B6/B10 hybrid offspring were intercrossed to generate mutants used for the mapping analysis. For each mutant sample, a panel of 123 B6/B10 polymorphic SNPs distributed throughout the mouse genome was PCR amplified, bar coded, and then pooled to generate a single library used for Ion Torrent sequencing. Sequencing carried out using the 314 chip yielded > 600,000 usable reads. These were aligned and mapped using a custom bioinformatics pipeline. Each SNP was sequenced to a depth > 500x, allowing accurate automated calling of the B6/B10 genotypes. This analysis mapped the mutation in Bishu to an interval on the proximal region of mouse chromosome 4. This was confirmed by parallel capillary sequencing of the 123 polymorphic SNPs. Further analysis of genes in the map interval identified a splicing mutation in Dnaic1 (c.204+1G > A), an intermediate chain dynein, as the disease causing mutation in Bishu. Overall, our experience shows Ion Torrent amplicon sequencing is high throughput and cost effective for conducting genome-wide mapping analysis and is easily scalable for other high volume genotyping analyses.

Item ID: 60712
Item Type: Article (Research - C1)
ISSN: 1432-1777
Keywords: Congenital Heart Disease; Primary Ciliary Dyskinesia; Amplicon Sequencing; Situs Inversus; Double Outlet Right Ventricle
Copyright Information: © Springer Science+Business Media New York 2013
Funders: National Institute of Health (NIH), USA
Projects and Grants: NIH Grants U01-HL098180, NIH Grants P30-HL101322
Date Deposited: 23 Oct 2019 12:36
FoR Codes: 06 BIOLOGICAL SCIENCES > 0604 Genetics > 060408 Genomics @ 30%
09 ENGINEERING > 0903 Biomedical Engineering > 090303 Biomedical Instrumentation @ 35%
08 INFORMATION AND COMPUTING SCIENCES > 0803 Computer Software > 080301 Bioinformatics Software @ 35%
SEO Codes: 92 HEALTH > 9201 Clinical Health (Organs, Diseases and Abnormal Conditions) > 920199 Clinical Health (Organs, Diseases and Abnormal Conditions) not elsewhere classified @ 100%
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