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Novel Sequence Discovery by Subtractive Genomics
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SNP discovery in non-model organisms using 454 next generation sequencing.

Christopher W Wheat1

  • 1Department of Biological and Environmental Sciences, University of Helsinki, Helsinki, Finland. chris@christopherwheat.net

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Summary

Developing single nucleotide polymorphisms (SNPs) from transcriptome sequencing is efficient for non-model species. This guide covers key challenges and methods for SNP discovery and development using RNA sequencing data.

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Area of Science:

  • Genomics
  • Bioinformatics
  • Molecular Biology

Background:

  • Roche 454 sequencing of the transcriptome is a standard method for obtaining single nucleotide polymorphisms (SNPs).
  • This approach is particularly valuable for non-model species lacking extensive genomic resources.

Purpose of the Study:

  • To present the primary issues and considerations for developing SNPs from transcriptome data in non-model species.
  • To compare transcriptome sequencing with RAD-tag sequencing for SNP discovery.
  • To discuss the potential of alternative sequencing platforms for SNP development.

Main Methods:

  • Detailed discussion of critical steps including tissue and sampling choices, mRNA preparation, normalization, pooling, and barcoding.
  • Guidance on determining sequencing depth, data assembly, and assembly assessment.
  • Methods for calling transcriptome SNPs and their conversion into genomic SNPs.

Main Results:

  • Identification of key challenges and best practices for transcriptome-based SNP discovery in non-model organisms.
  • Comparative analysis highlighting the strengths and weaknesses of transcriptome sequencing versus RAD-tag sequencing.
  • Evaluation of the utility of various sequencing platforms for SNP development.

Conclusions:

  • Transcriptome sequencing offers an efficient route to SNP discovery in non-model species.
  • Careful consideration of methodological choices is crucial for successful SNP development.
  • Further exploration of diverse sequencing technologies can enhance SNP discovery pipelines.