January 14-18, 2006
Town & Country Convention Center
San Diego, CA
John L. Norelli1 , Robert E. Farrell, Jr.2 , Carole L. Bassett1 , Angela M. Baldo3 , Herb S. Aldwinckle4 , Michael E. Wisniewski1
Fire blight, caused by the bacterium Erwinia amylovora, is a destructive disease of apple, pear and other plants in the subfamily Maloideae of the Rosaceae. The goal of this study was to use a global analysis of gene expression to characterize the temporal response of apple to infection by E. amylovora. 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 challenge and to obtain EST clones. By subtracting cDNAs synthesized from mRNAs expressed in one state (mock inoculated) from cDNAs derived from mRNAs expressed in another state (fire blight challenged) one can obtain sequences that are modulated when comparing the two mRNA populations side-by-side because sequences common to both populations are removed by hybridization. Gel electrophoresis of PCR-amplified subtracted cDNAs and unsubtracted controls indicated a greater quantity and size diversity in reverse subtracted samples (down-regulated ESTs) collected at 1 h and 2 h in comparison to forward subtracted samples (up-regulated) at 1 h, 2 h, 12 h, pooled early samples (15 min, 1 h, 2 h, 6 h, 12 h, and 24 h), and pooled later samples (48 h and 72 h); or in comparison to reverse subtracted samples (down-regulated) at 24 h and 48 h. PCR amplified subtracted cDNAs were cloned, sequenced, and identified by BLAST analysis. The project was supported by the National Research Initiative of the USDA Cooperative State Research, Education and Extension Service, grant number #2005-35300-15462.