1a.Objectives (from AD-416)
1. Identify MDV Meq and c-Jun binding sites in the chicken genome.
2. Identify differentially expressed genes following MDV-reactivation in MSB-1 cells.
3. Determine if differentially expressed genes identified in objective 2 also show differentially expression between MD tumors and CD4+ T cells.
4. Identify differentially expressed genes as a function of genetic resistance status or the presence of Meq.
1b.Approach (from AD-416)
1. Utilize chromatin immunoprecipitation (ChIP) using Meq or c-Jun antibodies and determine the DNA sequences via Solexa paired-end reads.
2. Isolate RNA from MSB-1 cells at time 0 and following MDV reactivation, and run on Affymetrix Chicken microarrays.
3. Compare by qRT-PCR the relative expression of key genes between MD tumors and CD4+ T cells.
4. Isolate RNA from line 6 (MD resistant) and line 7 (MD susceptible) CEF at various time before and after infection with either MDV or MDV lacking Meq, and run on Affymetrix Chicken microarrays.
This project is directly linked to specific agreement 3635-31320-008-26S “Positional Candidate Genes for Resistance to Marek’s Disease by Screening for Marek’s Disease Virus Meq-Regulated Genes.” The goal of this project is to identify chicken genes that confer resistance to Marek’s disease (MD), a significant issue to the poultry industry caused by the pathogenic Marek’s disease virus (MDV). MDV Meq is the likely oncogene and encodes a protein that controls expression of both viral and chicken genes. Using biotechnology, we are in the process of identifying regions in the chicken genome that Meq directly binds to and genes that are regulated by the presence of Meq. It is our hope that by combining these two sets of information we will be able to identify the genes controlled by Meq and the associated biological pathways. Thus far, we have DNA sequences for Meq-binding regions and are in the process of finding Meq-regulated genes. The project is proceeding as planned. This project is monitored by e-mail, telephone calls, and almost weekly face-to-face meeting between the two parties.