Abstract
BackgroundEven though microsatellite loci frequently have been isolated using recently developed next-generation sequencing (NGS) techniques, this task is still difficult because of the subsequent polymorphism screening requires a substantial amount of time. Selecting appropriate polymorphic microsatellites is a critical issue for ecological and evolutionary studies. However, the extent to which assembly strategy, read length, sequencing depth, and library layout produce a measurable effect on microsatellite marker development remains unclear. Here, we use six frog species for genome skimming and two frog species for transcriptome sequencing to develop microsatellite markers, and investigate the effect of different isolation strategies on the yield of microsatellites.ResultsThe results revealed that the number of isolated microsatellites increases with increased data quantity and read length. Assembly strategy could influence the yield and the polymorphism of microsatellite development. Larger k-mer sizes produced fewer total number of microsatellite loci, but these loci had a longer repeat length, suggesting greater polymorphism. However, the proportion of each type of nucleotide repeats was not affected; dinucleotide repeats were always the dominant type. Finally, the transcriptomic microsatellites displayed lower levels of polymorphisms and were less abundant than genomic microsatellites, but more likely to be functionally linked loci.ConclusionsThese observations provide deep insight into the evolution and distribution of microsatellites and how different isolation strategies affect microsatellite development using NGS.
Highlights
Even though microsatellite loci frequently have been isolated using recently developed next-generation sequencing (NGS) techniques, this task is still difficult because of the subsequent polymorphism screening requires a substantial amount of time
A total of 125.53 million raw PE reads were obtained from Illumina HiSeq (5.04 Gb and 7.32 Gb, for Amolops mantzorum and Quasipaa boulengeri, respectively)
In the era of NGS, Simple sequence repeats (SSRs) are still widely used in ecology and evolution [2], because they have their own advantages for population genetics compared to other molecular markers [55, 56]
Summary
Even though microsatellite loci frequently have been isolated using recently developed next-generation sequencing (NGS) techniques, this task is still difficult because of the subsequent polymorphism screening requires a substantial amount of time. For nonmodel species, microsatellites often have been developed de novo using this process, due to a low rate of cross-species microsatellite amplification success [5, 6] To overcome these obstacles, next-generation sequencing (NGS) has achieved faster and higher throughput nucleotide sequence data, which facilitate the identification and characterization of microsatellites in nonmodel species. The popularity of using NGS varies by platforms and by the sources of the sequences Both genome skimming and transcriptome sequencing have been used to mine microsatellite markers [18,19,20]
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