Copy Number Variation in the Horse Genome

We constructed a 400K WG tiling oligoarray for the horse and applied it for the discovery of copy number variations (CNVs) in 38 normal horses of 16 diverse breeds, and the Przewalski horse. Probes on the array represented 18,763 autosomal and X-linked genes, and intergenic, sub-telomeric and chrY sequences. We identified 258 CNV regions (CNVRs) across all autosomes, chrX and chrUn, but not in chrY. CNVs comprised 1.3% of the horse genome with chr12 being most enriched. American Miniature horses had the highest and American Quarter Horses the lowest number of CNVs in relation to Thoroughbred reference. The Przewalski horse was similar to native ponies and draft breeds. The majority of CNVRs involved genes, while 20% were located in intergenic regions. Similar to previous studies in horses and other mammals, molecular functions of CNV-associated genes were predominantly in sensory perception, immunity and reproduction. The findings were integrated with previous studies to generate a composite genome-wide dataset of 1476 CNVRs. Of these, 301 CNVRs were shared between studies, while 1174 were novel and require further validation. Integrated data revealed that to date, 41 out of over 400 breeds of the domestic horse have been analyzed for CNVs, of which 11 new breeds were added in this study. Finally, the composite CNV dataset was applied in a pilot study for the discovery of CNVs in 6 horses with XY disorders of sexual development. A homozygous deletion involving AKR1C gene cluster in chr29 in two affected horses was considered possibly causative because of the known role of AKR1C genes in testicular androgen synthesis and sexual development. While the findings improve and integrate the knowledge of CNVs in horses, they also show that for effective discovery of variants of biomedical importance, more breeds and individuals need to be analyzed using comparable methodological approaches.Sharmila Ghosh, Zhipeng Qu, Pranab J. Das, Erica Fang, Rytis Juras, E. Gus Cothran, Sue McDonell, Daniel G. Kenney, Teri L. Lear, David L. Adelson, Bhanu P. Chowdhary, Terje Raudsep

Chromosome-wise distribution of genic and intergenic CNVRs in the horse genome.

<p>Chromosome-wise distribution of genic and intergenic CNVRs in the horse genome.</p

Schematic of the homozygous deletion in chr29, 28.6–28.8 Mb in two XY DSD horses.

<p><b>A</b>. chr29 ideogram showing the location of <i>AKR1C</i> genes and a control gene <i>CREM</i>; <b>B</b>. Detailed map of the CNVR showing the location of genes (black horizontal bars) and CGH signal log2 values for 47 array probes in XY DSD and control horses; <b>C</b>. FISH results with a BAC 23N13 spanning the deletion (green signal) and a control BAC 76H13 for <i>CREM</i> from a non-CNVR (red signal); <b>D</b>. PCR with CNVR-specific primers in XY DSD and control horses.</p

Gene Ontology classifications of copy number variable genes in horses.

<p>Gene Ontology classifications of copy number variable genes in horses.</p

A CNVR map of the horse genome.

<p>Green line – loss; red line – gain; yellow line – complex; black dots – genes involved.</p

Novel and highly aberrant common (in bold font) CNVRs in XY DSD horses.

<p>Novel and highly aberrant common (in bold font) CNVRs in XY DSD horses.</p

Chromosome-wise CNVR statistics for the horse genome.

<p>Shared – found in 2 or more individuals; private – in one horse only; novel – not reported before; the horse genome statistics was retrieved from Ensembl (<a href="http://www.ensembl.org/index.html" target="_blank">http://www.ensembl.org/index.html</a>).</p><p>Chromosome-wise CNVR statistics for the horse genome.</p

Breed- and individual-wise summary of CNV calls in horses.

<p>The number of calls per individual was not significantly different (Student's T-test <i>p</i> = 0.07) between hair and blood DNA.</p><p>Breed- and individual-wise summary of CNV calls in horses.</p

A summary diagram of all CNV studies in horses and their contribution to the integrated CNV dataset.

<p>Numbers in arrow-heads denote the contribution of each study to the common pool of <b>301</b> shared CNVRs; numbers in arrow-tails denote the total and novel (separated by colon) CNVRs per study; CGH, SNP and NGS denote the platforms used for CNV detection.</p