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High-throughput sequencing and graph-based cluster analysis facilitate microsatellite development from a highly

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  • 1Department of Animal Behaviour Bielefeld University Postfach 100131 33501 Bielefeld Germany.

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|August 23, 2016
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Summary

Developing microsatellites in complex genomes is challenging. This study found that targeting unique DNA sequences, not repetitive elements, significantly improves microsatellite marker success in the club-legged grasshopper.

Keywords:
AcrididaeGomphocerus sibiricusOrthopteragenetic marker developmenthigh‐throughput sequencingmicrosatellitetransposable elements

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Area of Science:

  • Genomics
  • Molecular Ecology
  • Bioinformatics

Background:

  • Developing microsatellite markers for species with large, complex genomes is difficult.
  • Repetitive elements in genomes often interfere with microsatellite isolation.

Purpose of the Study:

  • To develop a novel approach for isolating polymorphic microsatellites from the club-legged grasshopper (Gomphocerus sibiricus).
  • To investigate the association between microsatellites and repetitive elements in complex genomes.

Main Methods:

  • Whole genome shotgun sequencing using Illumina MiSeq generated over three million paired-end reads.
  • RepeatExplorer software was used for graph-based clustering of repetitive DNA.
  • Primer pairs were designed from both clustered and singleton reads and tested for polymorphism.

Main Results:

  • Over 67% of sequencing reads were assigned to repetitive DNA clusters.
  • Primers designed from singleton reads were six times more successful in amplifying polymorphic microsatellite loci compared to those from clusters.
  • This indicates microsatellites associated with repetitive elements are less likely to amplify successfully.

Conclusions:

  • Microsatellite development in complex genomes can be improved by avoiding repetitive elements.
  • High-throughput sequencing combined with graph-based repetitive DNA analysis offers a viable strategy for isolating polymorphic microsatellites from challenging genomes.