Exome sequencing and arrayCGH detection of gene sequence and copy number variation between ILS and ISS mouse strains

Abstract

It has been well documented that genetic factors can influence predisposition to develop alcoholism. While the underlying genomic changes may be of several types, two of the most common and disease associated are copy number variations (CNVs) and sequence alterations of protein coding regions. The goal of this study was to identify CNVs and single-nucleotide polymorphisms that occur in gene coding regions that may play a role in influencing the risk of an individual developing alcoholism. Toward this end, two mouse strains were used that have been selectively bred based on their differential sensitivity to alcohol: the Inbred long sleep (ILS) and Inbred short sleep (ISS) mouse strains. Differences in initial response to alcohol have been linked to risk for alcoholism, and the ILS/ISS strains are used to investigate the genetics of initial sensitivity to alcohol. Array comparative genomic hybridization (arrayCGH) and exome sequencing were conducted to identify CNVs and gene coding sequence differences, respectively, between ILS and ISS mice. Mouse arrayCGH was performed using catalog Agilent 1 × 244 k mouse arrays. Subsequently, exome sequencing was carried out using an Illumina HiSeq 2000 instrument. ArrayCGH detected 74 CNVs that were strain-specific (38 ILS/36 ISS), including several ISS-specific deletions that contained genes implicated in brain function and neurotransmitter release. Among several interesting coding variations detected by exome sequencing was the gain of a premature stop codon in the alpha-amylase 2B (AMY2B) gene specifically in the ILS strain. In total, exome sequencing detected 2,597 and 1,768 strain-specific exonic gene variants in the ILS and ISS mice, respectively. This study represents the most comprehensive and detailed genomic comparison of ILS and ISS mouse strains to date. The two complementary genome-wide approaches identified strain-specific CNVs and gene coding sequence variations that should provide strong candidates to contribute to the alcohol-related phenotypic differences associated with these strains.

Genome characterization of the selected long- and short-sleep mouse lines

“…1b, c; Supplemental Figure S-2). By manual inspection, we found good correspondence to copy number variants (CNVs) that were previously identified by arrayCGH (Dumas et al 2014). A third of the previously called CNVs were confirmed (same call within ± 600 bp of breakpoints) using our pipeline.…”

“…We identified a large number of strain-specific sequence variants that can be used as markers to distinguish between the strains and, importantly, to help elucidate the genetic factors that contribute to the difference in acute alcohol sensitivity that exists between these strains. Previous work had identified ~40,000 SNPs (Saba et al 2011) and fewer than 100 SVs (Dumas et al 2014); here we report ~2.7 million SNPs and small indels, and over 7000 SVs that are different between the ILS and ISS, greatly expanding our knowledge of potential variants that contribute to phenotypic differences between the strains and among the LXS RIs. In addition, we have identified ~375,000 SNPs that are not present in dbSNP, implying that they have not previously been detected in any other sequenced mouse strains.…”

“…We also compared, by manual inspection, our unfiltered variants to a set of 7438 strain-specific variants identified by Dumas et al (2014) through exome sequencing. Most of these SNPs were also within our unfiltered list (98 % in ILS and 99 % in ISS).…”

Recurrent somatic loss of TNFRSF14 in classical Hodgkin lymphoma

“…Another limitation of our study is that insufficient numbers of flow sorted cells were available from patient cases to crossvalidate the results of genomic sequencing using conventional methods, such as FISH. Regardless, use of sequencing data to infer copy number states has been independently validated against conventional methods by several groups in a variety of contexts (Adey et al, ; Dumas et al, ; Liang et al, ; Wang et al, ). The various loci identified in the our study are supported by aCGH studies (Joos et al, ; Chui et al, ; Hartmann et al, ; Green et al, ; Steidl et al, ), including a deletion encompassing TNFRSF14 (Steidl et al, ), providing external support for the validity of our approach and findings.…”

Using the Coriell Personalized Medicine Collaborative Data to conduct a genome‐wide association study of sleep duration

“…Rs17122013 is located in the intronic region of Sortilin‐related VPS10 domain containing receptor 1 ( SORCS1 ) (Fig. ) which is deleted in an inbred short sleep mouse strain [Dumas et al, ]. Rs41463746 is located in the 3′ UTR of ELOVL fatty acid elongase 2 ( ELOVL2 ) (Fig.…”