Identification of defense-related rice genes by suppression subtractive hybridization and differential screening

Identification of host genes involved in defense responses is one of most c ritical steps leading to the elucidation of disease resistance mechanisms i n plants. In this study, two different cloning strategies were employed to identify defense-related genes from a tropical japonica rice cultivar (Oryz a sativa cv, Drew). With the use of bacterial colony arrays, differential s creening of a blast fungus (Pyricularia grisea)-induced rice cDNA library l ed to the isolation of 22 distinct rice genes that are expressed differenti ally in response to blast infection. Sequence analysis indicates that most of them are full-length cDNAs encoding pathogenesis-related proteins or oth er relatively abundant proteins. In combination with treatments of cyclohex imide plus jasmonic acid (JA) or benzothiadiazole (BTH) in rice seedlings, the polymerase chain reaction-based suppression subtractive hybridization a lso was conducted to search for immediate early (IE) defense-related genes whose transcription is independent of de novo protein synthesis. The initia l screening of only 768 subtracted clones resulted in the identification of 34 distinct IE genes that are induced by JA, BTH, and/or blast infection. Database searches revealed that these IE genes encode putative mitogen-acti vated protein kinase, diacylglycerol kinase, zinc finger protein, RelA-SpoT protein, ankyrin-containing protein, ABC transporter, beta -ketoacyl-CoA s ynthase, and other potential defense-signaling components. Further characte rization of these novel IE genes will likely facilitate the elucidation of defense signal transduction in rice plants.