Short Insertion and Deletion Discoveries via Whole-Genome Sequencing of 101 Thoroughbred Racehorses

Abstract

Thoroughbreds are some of the most famous racehorses worldwide and are currently animals of high economic value. To understand genomic variability in Thoroughbreds, we identified genome-wide insertions and deletions (INDELs) and obtained their allele frequencies in this study. INDELs were obtained from whole-genome sequencing data of 101 Thoroughbred racehorses by mapping sequence reads to the horse reference genome. By integrating individual data, 1,453,349 and 113,047 INDELs were identified in the autosomal (1–31) and X chromosomes, respectively, while 18 INDELs were identified on the mitochondrial genome, totaling 1,566,414 INDELs. Of those, 779,457 loci (49.8%) were novel INDELs, while 786,957 loci (50.2%) were already registered in Ensembl. The sizes of diallelic INDELs ranged from −286 to +476, and the majority, 717,736 (52.14%) and 220,672 (16.03%), were 1-bp and 2-bp variants, respectively. Numerous INDELs were found to have lower frequencies of alternative (Alt) alleles. Many rare variants with low Alt allele frequencies (<0.5%) were also detected. In addition, 5955 loci were genotyped as having a minor allele frequency of 0.5 and being heterogeneous genotypes in all the horses. While short-read sequencing and its mapping to reference genome is a simple way of detecting variants, fake variants may be detected. Therefore, our data help to identify true variants in Thoroughbred horses. The INDEL database we constructed will provide useful information for genetic studies and industrial applications in Thoroughbred horses, including a gene-editing test for gene-doping control and a parentage test using INDELs for horse registration and identification.

Construction of an individual identification panel for horses using insertion and deletion markers

“… *DB101: Allele frequency was extracted from a whole-genome variant database from 101 Thoroughbred horses (Tozaki et al ., 2023 [ 23 ]).PE: probability of exclusion, PD: power of discrimination, PID: probability of identity. …”

“…In our previous studies, we identified 12,173,068 SNVs and 1,566,396 INDELs in a population of 101 Thoroughbreds [ 22 , 23 ]. Approximately 7.77 times fewer INDELs were detected than SNVs, indicating that they are less likely to occur than single-base substitutions.…”

“…INDEL information was downloaded from DB101 (https://OSF.IO/QN45H/). DB101 is a database for INDELs that was constructed using the WGSs of 101 Thoroughbred horses in our previous study [ 23 ]. INDELs with reference allele frequencies in the range of 0.25–0.75 were extracted, and 39 INDELs were then selected based on chromosome and position information (non-neighbouring).…”

“…Through this procedure, whole-genome sequencing (WGS) of 88 horses (25 breeds) identified 23,559,582 SNVs and 2,396,022 insertions and deletions (INDELs) [ 8 ], and that of 534 horses (46 breeds) identified 29,038,030 SNVs and 2,102,379 INDELs [ 5 ]. In our previous studies, WGS of 101 Thoroughbred horses in Japan was analysed, and a whole-genome variant database, including loci and frequencies of approximately 12 million SNVs and 1.56 million short INDELs, was constructed [ 22 , 23 ]. This made it possible to search for diverse SNVs and INDELs.…”

“…Genotyping of urine extracts was performed four times, and the final genotype was confirmed synthetically (identical genotype with ≥2 markers). Finally,23,12,11, 28,6, and 20 markers were genotyped for BH, BP, BR, FP, LL, and PT, respectively.…”