Location: Southern Insect Management Research Unit
Project Number: 6066-22000-065-05
Start Date: Oct 01, 2008
End Date: Jul 31, 2011
As part of our preliminary work we have been able to develop a microarray containing 14,584 expressed sequence tags (EST) from H. virescens (see preliminary data). This array included 144 gut-specific ESTs obtained from a subtraction library. We have expanded the microarray by adding another 124 ESTs from a gut cDNA library. The first goal of aim 1 is to analyze expression of gut genes in susceptible and Cry1Ac-Cry2Ab resistant H. virescens larvae. We propose using a microarray in a high throughput analysis using mRNA labeled with CyDyes. To focus our analyses on proteins directly involved in resistance to both Cry1Ac and Cry2A toxins, we plan to compare susceptible (COW or ARS) with selected strains that are resistant to both toxins (CXC and KCBhyb for COW and EMS for ARS) and a strain that is only resistant to Cry1Ac (YHD2-B). Once the specific set of genes with altered expression is identified, we propose using quantitative real-time PCR (qRT-PCR) to confirm the differential expression levels in each strain. We expect this genomic approach to successfully identify genes whose products have a relevant role in the Cry intoxication process and whose alteration leads to resistance. These genes would be optimal candidates for the development of highly sensitive DNA-based resistance monitoring strategies.