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United States Department of Agriculture

Agricultural Research Service

Research Project: HIGH THROUGHPUT SEQUENCING FOR ASIAN SOYBEAN RUST AND PHYTOPHTHORA

Location: Corn Insects and Crop Genetics Research

2009 Annual Report


1a.Objectives (from AD-416)
The objectives are to identify differentially expressed genes in Rpp2 and rpp2 gentoypes, and Rps8 and rps8 genotypes during post-infection response to Asian Soybean Rust and Phytophthora Root and Stem Rot pathogens, respectively.


1b.Approach (from AD-416)
Soybean lines resistant (Rpp2) or susceptible (rpp2) to Asian Soybean Rust will be inoculated with the rust pathogen by collaborators in Brazil. Tissue samples will be collected at time zero, plus four other timepoints. RNA will be isolated from tissue representing each time point. Libraries will be made from each time point and provided to the cooperator. The cooperator will generate Solexa Digital Gene Expression data for each library. In addition, the cooperator will generate Solexa Whole Transcriptome Shotgun data from two pooled libraries representing resistant and sensitive lines. Soybean lines resistant (Rps8) or susceptible (rps8) to Phytophthora sojae will be inoculated with the Phytophthora pathogen. Tissue samples will be collected at time zero, plus three other timepoints. RNA will be isolated from tissue representing each time point. Libraries will be made from each timepoint and provided to the cooperator. The cooperator will generate Solexa Digital Gene Expression data for each library. In addition, the cooperator will generate Solexa Whole Transcriptome Shotgun data from libraries representing resistant and sensitive lines. The cooperator will generate a database of gene expression based on transcript counts and will provide this information to the ADODR. Differentially expressed genes will be overlain onto the soybean whole genome sequence and correlated with known positions of resistance genes.


3.Progress Report

The Ames group harvested roots from iron efficient Clark and Clark’s iron inefficient near-isogenic line. Root tissue was also harvested from T203, and A15. Tissue was from plants grown hydroponically in iron sufficient and iron insufficient conditions. Time points included one hour, 6 hours, and 24 hours post immersion in hydroponic solution. Leaf and root tissue from PI 399073 (Rps8), Williams, Williams 82, PI 86792-1, L83-570 (Rps3a), L76 1988 (Rps2) was harvested and RNA extracted. Leaf tissue from PI 547878 (Rpp2, Asian Soybean Rust resistance), PI 230970 (Rpp2), PI 437654, PI 548519 (BSR 101, Rps2) and L 82-2024 (Rps2) was harvested and RNA extracted. RNA from PI 547878 (Rpp2), PI 230970 (Rpp2), Williams, and Williams 82 was sent to collaborators at the National Center for Genome Resources for Solexa Digital Gene Expression sequencing to detect differentially expressed genes. Sequencing has been completed and data is currently being analyzed. Progress on this project is monitored through telephone calls, data exchange and written reports.


Last Modified: 9/1/2014
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