ARS | Publication request: Cloning and function validation of a nb-arc-lrr-type candidate gene for the greenbug aphid resistance locus, Gb3, in wheat

Submitted to: Plant and Animal Genome Conference
Publication Type: Abstract Only
Publication Acceptance Date: December 15, 2010
Publication Date: December 15, 2010
Citation: Azhaguvel, P., Weng, Y., Ma, Y., Luo, M., Simkova, H., Dolezel, J., Wicker, T., Saha, M., Rammna, H., Nelson, R., Zhou, C., Ray, T., Tang, Y., Liu, S., Rudd, J. 2010. Cloning and function validation of a nb-arc-lrr-type candidate gene for the greenbug aphid resistance locus, Gb3, in wheat [abstract]. Plant and Animal Genome Conference. P301.

Technical Abstract: The greenbug, Schizaphis graminum, is one of the most important aphid pests of small grain crops in many parts of the world. A single dominant gene, Gb3 originated from Aegilops tauschii has shown consistent and durable resistance against prevailing greenbug biotypes in wheat fields. A fine genetic map was developed for Gb3, which is located in the recombination-rich, telomeric bin of wheat chromosome arm 7DL. Markers closely linked with Gb3 were used to screen Ae. tauschii and ‘Chinese Spring’ 7DL–specific BAC libraries to initiate chromosome walking. Three clones were identified from the 7DL-specific BAC library and fully sequenced. Annotation of a total 373 kb DNA sequences identified a candidate gene for Gb3 which encodes a NB-ARC-LRR type R protein. Two resistant and susceptible Ae. tauschii accessions and one wheat line were sequenced in this candidate gene region and the full length cDNA sequences were also obtained to confirm the intron-exon splicing sites. The candidate gene consisted of three exons with a total length of 5,049 bp. Quantitative RT-PCR analysis revealed higher level transcription of this candidate gene in resistant lines than in susceptible lines before and after infestation of the greenbug. Barley stripe mosaic virus (BSMV)-based virus-induced gene silencing (VIGS) was employed to validate functions of the candidate gene..Four different region of the Gb3 candidate gene were cloned into BSMV VIGS vector for silencing. Plants inoculated with these VIGS constructs showed varying levels of reduction in endogenous transcript levels of the candidate gene. Aphid bioassay of VIGS-treated plants and genetic transformation of the candidate gene are underway.