17-24: The development of genetic engineering tools for cellulase producing fungus Acremonium cellulolyticus

Tuesday, May 1, 2012
Napoleon Ballroom C-D, 3rd fl (Sheraton New Orleans)
Tatsuya Fujii1, Hiroyuki Inoue1, Shinichi Yano1, Katsuji Murakami1 and Shigeki Sawayama2, (1)Biomass Technology Research Center, National Institute of Advanced Industrial Science and Technology, Higashi-Hiroshima, Japan, (2)Division of Applied Bioscience, Graduate School of Agriculture, Kyoto University, Kyoto, Japan
The filamentous fungi Acremonium cellulolyticus strain Y-94 is known as a cellulolytic and hemicellulolytic enzyme producer. In this study, we report for the first time, the whole genome sequence and analysis for strain Y-94. We obtained 60 scaffolds to generate 37.3 Mbp of genome sequences including 10,983 predicted ORFs. Among 10983 ORFs, 236 glycoside hydrolase family (GH) genes including 11 cellulolytic enzyme genes and 37 hemicellulolytic enzyme genes were found. We found 6 GH1 and 29 GH3 genes, which include b-glucosidase genes. Phylogenetic analyses revealed the possibility that A. cellulolytics Y-94 belongs to genus Penicillium.

Next, we attempted to the development of transformation system. We isolated a uracil auxotroph (strain CFP3) derived from strain CF-2612, which is a cellulase hyper-producing mutant originated from strain Y-94, and cloned a wild-type pyrF gene encoding orotate phosphoribosyl transferase (OPRTase) from Y-94. OPRTase activity was not detected in strain CFP3, which had one nucleotide substitution in its pyrF gene. The wild-type pyrF gene restored the defective growth of CFP3 on uracil-free medium, and PCR and Southern analyses revealed that wild-type pyrF was integrated into the genome. These results indicated that a transformation system for A. cellulolyticus with the pyrFgene as a selection marker was successfully developed.

These results will provide a set of instructions or suggestions about the improvements in cellulolytic enzyme production in A. cellulolyticus.

See more of: Poster Session 2
See more of: General Submissions