Genome Survey Sequencing, In-Silico and In-Vitro,] Development and Validation of Genomic Microsatellite Markers in Cooperatively Breeding Yellow-Billed Babbler ( Turdoides Affinis)

Abstract

Turdoides affinis is a species of group dwelling old world passerine of family Leiothrichidae. Unavailability of genome-wide sequence and species-specific molecular markers has hindered comprehensive understanding of phylogenetic and physiological aspects of T. affinis evolution. We generated genome-wide microsatellite markers through whole genome short read sequencing of T. affinis. A total of 195067054 reads were sequenced from 68.8 gigabytes of paired-end raw data, coverage 17X, genome size of 1.18 Gb, heterozygosity 0.97% and duplication rate of 0.97%. N50 value of SPAdes assembly was 14.2K and 44.7 % of all orthologous gene set in Passeriformes order was identified. 265297 microsatellites were mined covering around 0.49% of the genome. Of the 47 primer validated in-silico, 5 were randomly selected and in-vitro validated in 12 individuals of T. affinis. Genotyped data estimated observed heterozygosity (H₀) and expected heterozygosity (H_e) ratios range between 0.333 – 0.833 and 0.851 – 0.906 respectively, effective allele size ranged from 6.698 to 10.667, inbreeding coefficient of the population ranged from 0.080 – 0.631 and null allele frequency was calculated at 0.055 to 0.303. All the loci showed significant adherence to Hardy-Weinberg equilibrium. This study will facilitate future genomic studies on T. affinis and congeneric species.

Keywords: WGS, microsatellites, co-operative breeding, Turdoides affinis