Submitted to: International Congress on Molecular Plant-Microbe Interactions
Publication Type: Proceedings
Publication Acceptance Date: 5/6/2005
Publication Date: 12/12/2005
Citation: Norelli, J.L., Bassett, C.L., Farrell, R.E., Wisniewski, M.E. 2005. Rapid genomic response of malus to infection by erwinia amylovora. In: Proceedings of the International Congress on Molecular Plant-Microbe Interactions, December 14-19, 2005, Merida, Mexico. p. 190-191.
Technical Abstract: Plants in the Rosaceae subfamily Maloideae that are inoculated with Erwinia amylovora and incubated under favorable conditions will generally begin to develop the necrotic symptoms of fire blight disease within 3 to 7 days. cDNA subtractive hybridization was used to compare the populations of mRNA in mock inoculated (buffer controls) and E. amylovora inoculated "Gala" apple leaves at time intervals after inoculation and to obtain EST clones. Tissue samples were collected within 4 to 6 mm of inoculation site from both buffer and pathogen-challenged leaves. Total RNA was isolated from leaf samples, converted into cDNA, and used as tester (transcripts to be cloned after subtraction) and driver (reference transcripts removed) in both forward (pathogen inoculated = tester) and reverse (control = tester) cDNA hybridization subtraction. Gel electrophoresis of PCR amplified subtracted cDNAs and unsubtracted controls indicated a greater quantity and size diversity in reverse subtracted (down-regulated) samples collected at 2 h in comparison to early (15 min, 1 h, 2 h, 6 h, 12 h, and 24 h) and late (48 h and 72 h) forward subtracted (up-regulated), or 24 h and 48 h reverse subtracted samples. PCR amplified subtracted cDNAs were cloned, sequenced, and identified by BLAST analysis. ESTs of several photosynthesis related genes, as well as genes unrelated to photosynthesis, were identified in reverse cDNA hybridization subtractions 2 h after inoculation. Virtual Northern analysis confirmed that ribose-1,5 bisphosphate carboxylase small subunit (rbcS), chlorophyll a/b binding protein (CAB), a metallothionein-like protein (ATM2), and endomembrane protein 70 (BAB01926) were down-regulated 2 h following inoculation. Expression of CAB and BAB01926 were equivalent to controls 6 h and 12 h after inoculation.