A pipeline for effectively developing highly polymorphic simple sequence repeats markers based on multi‐sample genomic data

Abstract

Simple sequence repeats (SSRs) are widely used genetic markers in ecology, evolution, and conservation even in the genomics era, while a general limitation to their application is the difficulty of developing polymorphic SSR markers. Next‐generation sequencing (NGS) offers the opportunity for the rapid development of SSRs; however, previous studies developing SSRs using genomic data from only one individual need redundant experiments to test the polymorphisms of SSRs. In this study, we designed a pipeline for the rapid development of polymorphic SSR markers from multi‐sample genomic data. We used bioinformatic software to genotype multiple individuals using resequencing data, detected highly polymorphic SSRs prior to experimental validation, significantly improved the efficiency and reduced the experimental effort. The pipeline was successfully applied to a globally threatened species, the brown eared‐pheasant (Crossoptilon mantchuricum), which showed very low genomic diversity. The 20 newly developed SSR markers were highly polymorphic, the average number of alleles was much higher than the genomic average. We also evaluated the effect of the number of individuals and sequencing depth on the SSR mining results, and we found that 10 individuals and ~10X sequencing data were enough to obtain a sufficient number of polymorphic SSRs, even for species with low genetic diversity. Furthermore, the genome assembly of NGS data from the optimal number of individuals and sequencing depth can be used as an alternative reference genome if a high‐quality genome is not available. Our pipeline provided a paradigm for the application of NGS technology to mining and developing molecular markers for ecological and evolutionary studies.

Genetic Characterization of Endangered Indian Mithun (Bos frontalis), Indian Bison/Wild Gaur (Bos gaurus) and Tho-tho Cattle (Bos indicus) Populations Using SSR Markers Reveals Their Diversity and Unique Phylogenetic Status

“…SSRs have been the most frequently used genetic marker in population genetics in the past two decades [23]. Recently, Single Nucleotide Polymorphic loci (SNPs) have been used in a variety of genetic studies [24], but they could not replace SSRs completely, even in the genomic era, due to their potential benefits [25]. Multi-allelic nature and high polymorphism of SSR markers help to establish the relationship among the individuals even with fewer markers.…”

Microsatellite Genome-Wide Database Development for the Commercial Blackhead Seabream (Acanthopagrus schlegelii)

“…Bhattarai et al [ 37 ] identified a final set of the 5986 polymorphic SSR loci using the HipSTR program by comparing and counting repeat number variation across the SSR loci of the whole-genome sequence data (30×) among 21 spinach plants. A total of 12,549 SSR loci were genotyped successfully from resequencing data (20×) of 20 brown-eared pheasant individuals using lobSTR software [ 38 ]. Eight hundred and seventy-one polymorphic SSRs were screened across the transcriptome of twelve individuals using the PSR tool in swamp eel [ 30 ].…”

Genome-wide survey and characterization of microsatellites in cashew and design of a web-based microsatellite database: CMDB

“…The NGS technologies allow rapid large-scale sequencings at a lesser cost, which permits discovery and development of SSR markers for less studied crops ( Khodaeiaminjan et al., 2018 ; Taheri et al., 2018 ; Luo et al., 2021 ; Wang et al., 2022 ). The genome sequences generated using the NGS technology have been used to discover genome-wide SSRs and develop SSR markers in various tree species, such as pistachio ( Ziya Motalebipour et al., 2016 ), hazelnut ( Öztürk et al., 2018 ), avocado ( Ge et al., 2019 ), fruit and forest species ( Song et al., 2021 ), and Grevillea sp.…”

“…Previously, the development of microsatellite or SSR markers was an expensive and time-consuming process. But the rapid improvements in sequencing technologies and bioinformatics have reduced the cost and time required for the development of a large set of robust markers through genome sequencing and in silico mining of the genome sequences for potential markers ( Luo et al., 2021 ; Wang et al., 2022 ; Savadi et al., 2023 ). To date, there have been no studies on the genome-wide characterization of microsatellites or SSRs in cashew.…”