De novo assembly of transcriptome from next-generation sequencing data

Abstract

Reconstruction of transcriptome by de novo assembly from next generation sequencing (NGS) short-sequence reads provides an essential mean to catalog expressed genes, identify splicing isoforms, and capture the expression detail of transcripts for organisms with no reference genome available. De novo transcriptome assembly faces many unique challenges, including alternative splicing, variable expression level covering a dynamic range of several orders of magnitude, artifacts introduced by reverse transcription, etc. In the current review, we illustrate the grand strategy in applying De Bruijn Graph (DBG) approach in de novo transcriptome assembly. We further analyze many parameters proven critical in transcriptome assembly using DBG. Among them, k-mer length, coverage depth of reads, genome complexity, performance of different programs are addressed in greater details. A multi-k-mer strategy balancing efficiency and sensitivity is discussed and highly recommended for de novo transcriptome assembly. Future direction points to the combination of NGS and third generation sequencing technology that would greatly enhance the power of de novo transcriptomics study.

Graphical abstract

Keywords

transcriptome / de novo assembly / De Bruijn Graph / next generation sequencing / k-mer length / RNA splicing / performance

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ABBREVIATIONS

CAGE, cap analysis of gene expression; cDNA, complementary DNA; CDS, coding sequence; CPU, central processing unit; DBG, De Bruijn Graph; MPSS, massively parallel signature sequencing; NGS, next generation sequencing; ORF, open reading frame; RMBT, reads mapped back to assembled transcripts; SAGE, serial analysis of gene expression; SNV, single nucleotide variation; SOLiD, sequencing by oligonucleotide ligation and detection; UTR, untranslated region

ACKNOWLEDGEMENTS

This work is supported in part by grants from the National Basic Research Program of China (Nos. 2012CB316501 and 2013CB127000) and the National Natural Science Foundation of China (Nos. 31571310 and 31271409).

COMPLIANCE WITH ETHICS GUIDELINES

The authors Xuan Li, Yimeng Kong, Qiong-Yi Zhao, Yuan-Yuan Li and Pei Hao declare they have no conflict of interests.

This article does not contain any studies with human or animal subjects performed by any of the authors.

Funding

RIGHTS & PERMISSIONS

2016 Higher Education Press and Springer-Verlag Berlin Heidelberg