A simple hybridization-based strategy for the generation of non-redundant EST collections—a case study in barley (Hordeum vulgare L.)

R.K. Varshney; H. Zhang; E. Potokina; N. Stein; P. Langridge; A. Graner

doi:10.1016/j.plantsci.2004.05.011

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A simple hybridization-based strategy for the generation of non-redundant EST collections—a case study in barley (Hordeum vulgare L.)

Journal article

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A simple hybridization-based strategy for the generation of non-redundant EST collections—a case study in barley (Hordeum vulgare L.)

R.K. Varshney, H. Zhang, E. Potokina, N. Stein, P. Langridge and A. Graner

Plant Science, Vol.167(3), pp.629-634

2004

DOI: https://doi.org/10.1016/j.plantsci.2004.05.011

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Abstract

The systematic development of functional molecular markers requires a set of non-redundant ESTs. However large-scale EST sequencing projects are frequently based on random sequencing of shotgun cDNA clones, which usually results in high redundancy of ESTs. To increase the efficiency in the identification of novel ESTs/genes for the development of non-redundant markers, it is necessary to reduce the redundancy of sequencing by eliminating the redundant clones from the cDNA library. In order to reach this goal, we adopted the ‘total cDNA hybridization’ approach on a set of barley cDNAs from different libraries to remove redundant clones prior to sequencing. To quantify the accuracy of the approach we performed cluster analysis on the ESTs of the clones used in this study. The results show that up to 90% of the redundant clones that are present in clusters can be identified by this approach. Furthermore cluster analysis revealed that the set of non-hybridizing cDNA clones contains 82–89% ‘singletons’. Therefore, cDNA hybridization of arrayed colony filters represents an efficient and cheap procedure to deplete a cDNA library of redundant clones in order to generate non-redundant molecular markers. In addition, this approach is suitable for maximizing the rate of gene discovery in an EST programme.

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Title: A simple hybridization-based strategy for the generation of non-redundant EST collections—a case study in barley (Hordeum vulgare L.)
Authors/Creators: R.K. Varshney (Author/Creator) - Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK)
H. Zhang (Author/Creator) - Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK)
E. Potokina (Author/Creator) - Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK)
N. Stein (Author/Creator) - Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK)
P. Langridge (Author/Creator) - Australian Centre for Plant Functional Genomics
A. Graner (Author/Creator) - Leibniz-Institut für Pflanzengenetik und Kulturpflanzenforschung (IPK)
Publication Details: Plant Science, Vol.167(3), pp.629-634
Publisher: Elsevier
Identifiers: 991005546369507891
Copyright: © 2004 Elsevier Ireland Ltd.
Murdoch Affiliation: Murdoch University
Language: English
Resource Type: Journal article

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Collaboration types: Domestic collaboration; International collaboration
Citation topics: 3 Agriculture, Environment & Ecology; 3.4 Crop Science; 3.4.96 QTL
Web Of Science research areas: Biochemistry & Molecular Biology; Plant Sciences
ESI research areas: Plant & Animal Science