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Open Access Highly Accessed Research article

Analysis of 4,664 high-quality sequence-finished poplar full-length cDNA clones and their utility for the discovery of genes responding to insect feeding

Steven G Ralph16, Hye Jung E Chun2, Dawn Cooper1, Robert Kirkpatrick2, Natalia Kolosova13, Lee Gunter4, Gerald A Tuskan4, Carl J Douglas3, Robert A Holt2, Steven JM Jones2, Marco A Marra2 and Jörg Bohlmann135*

Author Affiliations

1 Michael Smith Laboratories, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada

2 British Columbia Cancer Agency Genome Sciences Centre, Vancouver, British Columbia, V5Z 4E6, Canada

3 Department of Botany, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada

4 Environmental Sciences Division, Oak Ridge National Laboratory, Oak Ridge, Tennessee, 37831, USA

5 Department of Forest Sciences, University of British Columbia, Vancouver, British Columbia, V6T 1Z4, Canada

6 Department of Biology, University of North Dakota, Grand Forks, North Dakota, 58202-9019, USA

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BMC Genomics 2008, 9:57  doi:10.1186/1471-2164-9-57

Published: 29 January 2008

Abstract

Background

The genus Populus includes poplars, aspens and cottonwoods, which will be collectively referred to as poplars hereafter unless otherwise specified. Poplars are the dominant tree species in many forest ecosystems in the Northern Hemisphere and are of substantial economic value in plantation forestry. Poplar has been established as a model system for genomics studies of growth, development, and adaptation of woody perennial plants including secondary xylem formation, dormancy, adaptation to local environments, and biotic interactions.

Results

As part of the poplar genome sequencing project and the development of genomic resources for poplar, we have generated a full-length (FL)-cDNA collection using the biotinylated CAP trapper method. We constructed four FLcDNA libraries using RNA from xylem, phloem and cambium, and green shoot tips and leaves from the P. trichocarpa Nisqually-1 genotype, as well as insect-attacked leaves of the P. trichocarpa × P. deltoides hybrid. Following careful selection of candidate cDNA clones, we used a combined strategy of paired end reads and primer walking to generate a set of 4,664 high-accuracy, sequence-verified FLcDNAs, which clustered into 3,990 putative unique genes. Mapping FLcDNAs to the poplar genome sequence combined with BLAST comparisons to previously predicted protein coding sequences in the poplar genome identified 39 FLcDNAs that likely localize to gaps in the current genome sequence assembly. Another 173 FLcDNAs mapped to the genome sequence but were not included among the previously predicted genes in the poplar genome. Comparative sequence analysis against Arabidopsis thaliana and other species in the non-redundant database of GenBank revealed that 11.5% of the poplar FLcDNAs display no significant sequence similarity to other plant proteins. By mapping the poplar FLcDNAs against transcriptome data previously obtained with a 15.5 K cDNA microarray, we identified 153 FLcDNA clones for genes that were differentially expressed in poplar leaves attacked by forest tent caterpillars.

Conclusion

This study has generated a high-quality FLcDNA resource for poplar and the third largest FLcDNA collection published to date for any plant species. We successfully used the FLcDNA sequences to reassess gene prediction in the poplar genome sequence, perform comparative sequence annotation, and identify differentially expressed transcripts associated with defense against insects. The FLcDNA sequences will be essential to the ongoing curation and annotation of the poplar genome, in particular for targeting gaps in the current genome assembly and further improvement of gene predictions. The physical FLcDNA clones will serve as useful reagents for functional genomics research in areas such as analysis of gene functions in defense against insects and perennial growth. Sequences from this study have been deposited in NCBI GenBank under the accession numbers EF144175 to EF148838.