Project Number: 6618-22000-039-27
Project Type: General Cooperative Agreement

Start Date: May 01, 2012
End Date: Apr 26, 2013

Objective:
1. To generate transcriptome profiles of both susceptible and resistant citrus responding to huanglongbing (HLB) infection using RNA-Seq technology. 2. To identify key resistant genes from differentially expressed genes and gene clusters between the HLB-susceptible and HLB-resistant plants via intensive bioinformatics and other experimental verifications such as real-time polymerase chain reaction (RT-PCR). 3. To create transgenic citrus cultivars with new constructs containing the resistant gene(s).

Approach:
1. Extract mRNA from 4 susceptible and 4 resistant plants from each of three progenies (24 genotypes) and use next generation sequencing technology to sequence all mRNA. 2. Clean the reads from RNA-Seq and assemble the RNA-Seq reads to generate transcriptome. 3. Identify differentially expressed genes using statistical tests. 4. Annotate differentially expressed genes. 5. Use random matrix theory-based method to identify differentially expressed gene networks. 6. Use real time polymerase chain reaction (PCR) to verify the differentially expressed genes in original 24 genotypes. Consistency of these resistance/susceptibility genes will be further verified through evaluation of 24 additional highly resistant and susceptible genotypes from more diverse genetic backgrounds. 7. Most promising genes will be used in citrus-only constructs and used to create transgenic citrus.