Development of genomic microsatellite markers in cluster bean using next-generation DNA sequencing and their utility in diversity analysis

Abstract

With high global demand, seeds and gum of cluster bean are an essential raw material for various industries. Worldwide the low yield and productivity of guar (Cyamopsis tetragonoloba) create a large gap between demand and supply of guar. Therefore, to upsurge guar production, there is a need to improve the guar at genetic level. The genetic improvement of cluster bean is slower due to insufficient genomic resources, co-dominant marker system especially simple sequence repeats (SSRs) and inadequate information on the variability of germplasm. For development of microsatellite markers in cluster bean, DNA survey sequencing was carried out using the Miseq NGS system. DNA sequencing generated 73,934 sequences which harboured microsatellite repeat sequences. Successfully, a total of 15,399 marker-pairs were designed. The amplicon size ranged between 101–385 base-pairs. To validate the primers and to analyse the diversity in released varieties and germplasm of cluster bean, successfully amplifiable 21 primers pairs were used. With a mean of 2.05 alleles per primers, 21 bands were detected. The polymorphism information content (PIC) for polymorphic markers fluctuated between 0.04 - 0.67 (mean = 0.3). With the average dissimilarity coefficient of 0.11, four clusters were noticed with NJ analysis. The grouping pattern of genotypes indicated a low genetic variability in the guar gene pool. The results of this study suggested that instead of re-circulation of parental lines, hybrid breeding using diverse parents followed by selection should be performed for genetic improvement of cluster bean. Keywords: Cluster bean, Diversity, DNA marker, NGS, SSR