A pipeline for the de novo assembly of the Themira biloba (Sepsidae: Diptera) transcriptome using a multiple k-mer length approach

BMC Genomics

Julia H Bowsher¹  Ian Dworkin²  Alex S Torson¹  Dacotah Melicher¹ 

[1] Department of Biological Sciences, North Dakota State University, 1340 Bolley Drive, 218 Stevens Hall, Fargo, ND 58102, USA;Department of Zoology, Michigan State University, 328 Giltner Hall, East Lansing, MI 48823, USA

关键词: Cloud computing;    Pipeline;    Transcriptome;    Sepsidae;    de novo assembly;    Multiple k-mer;   

Others  :  1217771
DOI  :  10.1186/1471-2164-15-188

 received in 2013-11-04, accepted in 2014-03-03,  发布年份 2014

【 摘 要 】

Background

The Sepsidae family of flies is a model for investigating how sexual selection shapes courtship and sexual dimorphism in a comparative framework. However, like many non-model systems, there are few molecular resources available. Large-scale sequencing and assembly have not been performed in any sepsid, and the lack of a closely related genome makes investigation of gene expression challenging. Our goal was to develop an automated pipeline for de novo transcriptome assembly, and to use that pipeline to assemble and analyze the transcriptome of the sepsid Themira biloba.

Results

Our bioinformatics pipeline uses cloud computing services to assemble and analyze the transcriptome with off-site data management, processing, and backup. It uses a multiple k-mer length approach combined with a second meta-assembly to extend transcripts and recover more bases of transcript sequences than standard single k-mer assembly. We used 454 sequencing to generate 1.48 million reads from cDNA generated from embryo, larva, and pupae of T. biloba and assembled a transcriptome consisting of 24,495 contigs. Annotation identified 16,705 transcripts, including those involved in embryogenesis and limb patterning. We assembled transcriptomes from an additional three non-model organisms to demonstrate that our pipeline assembled a higher-quality transcriptome than single k-mer approaches across multiple species.

Conclusions

The pipeline we have developed for assembly and analysis increases contig length, recovers unique transcripts, and assembles more base pairs than other methods through the use of a meta-assembly. The T. biloba transcriptome is a critical resource for performing large-scale RNA-Seq investigations of gene expression patterns, and is the first transcriptome sequenced in this Dipteran family.

【 授权许可】

   
2014 Melicher et al.; licensee BioMed Central Ltd.

【 图 表 】

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