Introduction marker gene serin Green Fluorescent Protein (sGFP) to Pyricularia grisea Rases 001 and 173 using Agrobacterium tumefaciens

Abstract

Pyricularia grisea (teleomorf: Magnaporthe grisea) is causal agent of rice blast disease. Dominant races of P. grisea attacking rice in Indonesia is 001 and 173 races. Race 001 have low level of virulence but the ability to survive in the field is very high, while the 173 race have high level of virulence, but the ability to survive in field is low. GFP gene has been successfully used as amonitor for a blast disease caused by P. grisea. Therefore, GFP gene could be used to monitor the pathogenesis of both races. Introduction of serin GFP (sGFP) gene into spores of P. grisea by using A. tumefaciens- mediated transformation. sGFP gene has been introduced into A. tumefaciens LBA 4404 by triparental mating. Pyricularia grisea-sGFP was selected using selection medium that contains 300 μg/ml hygromycin. Transformation results shown P. grisea 001 races-sGFP transformant and P. grisea 173 races-sGFP transformant were not different with non-transformant on growth rate. To determine the insertion of sGFP gene into genome of P.grisea was detected by using PCR method with specific primer for sGFP. The result shown that amplification of sGFP gene was successfully and thus indicated that P.grisea 001 races-sGFP and 173 races-sGFP contained sGFP gene.