Metabolomics,Unknown,Transcriptomics,Genomics,Proteomics

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Lr1-mediated leaf rust resistance pathways of transgenic wheat lines

ABSTRACT: The wheat gene Lr1 imparts resistance to certain races of the leaf rust pathogen Puccinia triticina. However, the biological basis underlying the complex resistance mechanisms remains largely unknown. Gene expression profiling analyses using the Affymetrix GeneChipM-BM-. Wheat Genome Array were performed with six independent Lr1 transgenic lines sampled before inoculation (0h) and at 6 and 24 h after inoculation with the avirulent P. triticina race CCDS. For the 6 and 24 h time points, 3M-BM- 384 probe sets corresponding to 2M-BM- 978 transcripts were differentially expressed as compared to the plants before inoculation. Genes involved in defence signalling were triggered as early as 6 h post inoculation (HPI) and included COI1, the primary component of jasmonic acid signalling. By 24 HPI, a shift in plant cellular metabolism, namely carbon conservation, was notable. The enzymes of glycolysis and gluconeogenesis were down-regulated, however, enzymes linked to alternative carbon sequestration processes such as the glyoxylate cycle were up-regulated. Also at 24 HPI, various secondary signalling molecules belonging to the various hormone signalling pathways were triggered. These microarray data suggest that the Lr1 mediated leaf rust resistance is achieved by early onset of defence signalling followed by co-ordinated interplay of metabolism alteration and elicitation of defence responses. We used microarrays to detail the early gene expression events underlying Lr1-mediated resistance to Puccinia triticina pathogen in wheat. Three biological replicates with six independent Lr1 transgenic lines were used in this study to eliminate the possibility of false discovery of genes due to copy number variation or the position of insertion of the transgene Lr1. Early time points of 6 and 24 HPI were compared to 0 h (prior to inoculation) to capture the early signalling events involved in Lr1-mediated response to rust infection.

ORGANISM(S): Triticum aestivum

SUBMITTER: Sylvie Cloutier

PROVIDER: E-GEOD-32151 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress

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Project description:Transcriptional changes were monitored in the wheat cultivar Avocet*6/Yr1 following inoculation with avirulent and virulent wheat yellow rust (Puccinia striiformis f.sp. tritici) isolates using the Affymetrix wheat genome array GeneChip®. Seedlings of Avocet*6/Yr1 were grown to growth stage 12-13 (Zadoks et al., 1974) before inoculating with either the P.s. f.sp. tritici isolate 169E136 (Yr1-virulent) or 232E137 (Yr1-avirulent) or with talc for the Mock inoculation (Boyd and Minchin 2001). Plants were grown under a 16/8 hour photoperiod cycle, supplemented with sodium lighting (240 ?mol m-2 s-1) at day/night temperatures of 20°C/15°C and a relative humidity of 60%, both before and after inoculation, in a spore-free, containment level 2 greenhouse. Leaf samples from six seedlings were collected 6, 12, 24, 48 and 72 hpi for RNA extraction and transcriptomics analysis from three independent biological experiments. Leaf tissue was ground under liquid nitrogen and total RNA extracted using the TRIzol reagent method, following the manufacturer’s instructions (Invitrogen, Carlsbad, CA). RNA from each time point (16 ?g) were pooled to obtain 80 ?g of total RNA for each treatment (mock inoculated control; avirulent isolate inoculated; virulent isolate inoculated). Pooled RNA samples were further purified using the Qiagen RNeasy mini-kit according to the manufacturer instructions (Qiagen) and RNA integrity was confirmed using the Agilent 2100 Bioanalyzer (Agilent). Affymetrix GeneChip processing, including RNA quality control, microarray hybridisation and data acquisition was performed through contract research services by the John Innes Genome Laboratory, John Innes Centre, Norwich, U.K. A total of nine hybridisations were performed. ****[PLEXdb(http://www.plexdb.org) has submitted this series at GEO on behalf of the original contributor, Osman Bozkurt. The equivalent experiment is TA25 at PLEXdb.] pathogen isolates: Mock inoculated (Control)(3-replications); pathogen isolates: Yr1-avirulent wheat yellow rust isolate 232E137(3-replications); pathogen isolates: Yr1-virulent wheat yellow rust isolate 169E136(3-replications)

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Lr1-mediated leaf rust resistance pathways of transgenic wheat lines

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