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Title: Laser capture microdissection and expressed sequence tag analysis of uredinia formed by Phakopsora pachyrhizi, the causal agent of Asian soybean rust

Author
item Tremblay, Arianne
item Li, Shuxian
item Scheffler, Brian
item Matthews, Benjamin

Submitted to: Physiological and Molecular Plant Pathology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 6/9/2009
Publication Date: 9/24/2009
Citation: Tremblay, A., Li, S., Scheffler, B.E., Matthews, B.F. 2009. Laser capture microdissection (LCM) and expressed sequence tag analysis of uredinia formed by Phakopsora pachyrhizi, the causal agent of Asian soybean rust. Physiological and Molecular Plant Pathology. 73(6):163-174.

Interpretive Summary: Asian soybean rust is a devastating fungal pathogen of a wide range of host plants. It arrived in the U.S. in 2004 and is a threat to the soybean industry. Little is known about which genes are expressed during the life cycle of Asian soybean rust. However this knowledge is important, if scientists are to develop molecular approaches to broadening resistance of soybean to the various isolates of the rust. Through DNA sequencing and analysis we identified genes of Asian soybean rust that are expressed by the fungus when it is mature and developing new spores for the next generation. If scientists can develop a method to inhibit rust sporulation, then the rust cannot reproduce and the disease cannot spread to other plants. Some of these fungal genes will be useful to scientists interested in developing methods to inhibit the attack and spread of Asian soybean rust.

Technical Abstract: Soybean is one of the top five agricultural products in the U.S. Protection of soybean from present and new exotic pathogens is very important for soybean production. Soybean rust is caused by the obligate fungus Phakopsora pachyrhizi Sydow, an exotic pathogen. This pathogen causes yield losses due to premature defoliation, fewer seeds per pod and decreased number of filled pods per plant. From this perspective we identified genes from P. pachyrhizi that might be involved in the infection process specifically during the uredinium formation. Thus, we constructed and analyzed cDNA library to identify candidate genes. The library was constructed from RNA isolated from uredinium formed by P. pachyrhizi on the under side of leaves. Uredinia were isolated by laser capture microdissection. A portion of the library was sequenced, contigs were formed, and blast searches were conducted to determine the identity of the genes. We found 41 expressed sequence tags (EST) with significant similarities to (Evalue< 10-5) to sequences deposited in the NCBI non-redundant protein database. Of those ESTs, 61% was similar to fungal sequences. Even if 59% of those significant ESTs shown identities with hypothetical proteins or proteins with unknown function, some shown identities with proteins involved in energy production, cellular communication/signal transduction, and transcription. RT-PCR was used to confirm expression of six rust genes. In the future, target pathogen genes will be studied to determine if they can be used to control ASR in soybean.