Transcriptome-scale homoeolog-specific transcript assemblies of bread wheat

Andreas W Schreiber,Ute Baumann,Kerrie L Forrest,Stephan L Kong,Matthew J Hayden,Peter Langridge

doi:10.1186/1471-2164-13-492

Abstract

BackgroundBread wheat is one of the world’s most important food crops and considerable efforts have been made to develop genomic resources for this species. This includes an on-going project by the International Wheat Genome Sequencing Consortium to assemble its large and complex genome, which is hexaploid and contains three closely related ‘homoeologous’ copies for each chromosome. This multi-national effort avoids the complications polyploidy entails for correct assembly of the genome by sequencing flow-sorted chromosome arms one at a time. Here we report on an alternate approach, a direct homoeolog-specific assembly of the expressed portion of the genome, the transcriptome.ResultsAfter assessment of the ability of various assemblers to generate homoeolog-specific assemblies, we employed a two-stage assembly process to produce a high-quality assembly of the transcriptome of hexaploid wheat from Roche-454 and Illumina GAIIx paired-end sequence reads. The assembly process made use of a rapid partitioning of expressed sequences into homoeologous clusters, followed by a parallel high-fidelity assembly of each cluster on a 1150-processor compute cloud. We assessed assembly quality through comparison to known wheat gene sequences and found that in ca. 98.5% of cases the assembly was sufficiently accurate for homoeologous triplets to be cleanly separated into either two or three separate contigs. Comparison to publicly available transcript collections suggests that the assembly covers ~75-80% of the complete transcriptome.ConclusionsThis work therefore describes the first homoeolog-specific sequence assembly of the wheat transcriptome and provides a reference transcriptome for future wheat research. Furthermore, our assembly methodology is transferable to other polyploid organisms.

Highlights

Bread wheat is one of the world’s most important food crops and considerable efforts have been made to develop genomic resources for this species
We describe our work towards the sequencing and subsequent homoeolog-specific assembly of the wheat transcriptome and show, through comparison with existing sequence resources, that the resultant assembly goes a long way towards producing a comprehensive compendium of the gene sequences of bread wheat
Sequencing of the wheat transcriptome Wheat mRNA from a single cultivar, the elite variety “Kukri” [22,23], was sequenced using: a) short-read Illumina GAIIx technology for sequencing depth, and b) long-read Roche GSFLX Titanium technology for homoeolog-sensitivity

Summary

Introduction

Bread wheat is one of the world’s most important food crops and considerable efforts have been made to develop genomic resources for this species This includes an on-going project by the International Wheat Genome Sequencing Consortium to assemble its large and complex genome, which is hexaploid and contains three closely related ‘homoeologous’ copies for each chromosome. This multi-national effort avoids the complications polyploidy entails for correct assembly of the genome by sequencing flow-sorted chromosome arms one at a time. Any computational procedure for assembling such large and complex genomes must, be exceedingly efficient with both time and memory resources, but at the same time must be highly accurate to avoid mis-assembly of closely related sequences For this reason, the sequencing of bread wheat

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