Polymorphic microsatellite loci from ddRAD sequencing data in different Indian yak populations adapted to diverse climatic conditions

Microsatellites are highly polymorphic, codominant, and reproducible molecular markers that are fairly evenly distributed across the eukaryotic genomes. Although cattle-specific microsatellite markers have been employed in yak, no species-specific marker set has yet been established in this unique species. The present study aimed to identify and validate polymorphic microsatellite markers in four distinct Indian yak cohorts, i.e., Arunachali, Ladakhi, Himachali, and Sikkimi using double-digest restriction-site associated DNA (ddRAD) sequencing data from 20 representative animals per cohort (n = 80). Approximately 260 million raw paired-end reads were retrieved and processed in the present study. Paired-end reads (n = 40 per cohort) were merged using PEAR program, and the assembled reads were screened for RAD cut sites. Polymorphic microsatellite loci were elucidated and corresponding primer sets were designed using QDD program, utilizing only unique consensus sequences and singletons for primer designing. In-silico PCR amplification was performed using in_silico_PCR.pl, implemented in Perl to validate primer specificity. A total of 32, 82, 149, and 31 polymorphic SSR loci were identified in Sikkimi, Himachali, Arunachali, and Ladakhi cohorts, respectively; of which 26, 72, 106, and 24 loci were successfully validated through in-silico amplification. A total of 32 microsatellites were common between the Himachali and Arunachali cohorts, while two loci were shared across all populations. On the other hand, 13, 39, 56, and eight loci were private to Sikkimi, Himachali, Arunachali, and Ladakhi yak, respectively. The validated polymorphic SSR markers provide a valuable genomic resource for assessing genetic diversity, population structure, and evolutionary adaptation in yak populations, and can be further evaluated and validated through wet-laboratory experimentation on larger cohorts.