关键词:

基因敲除, fluG基因, 产孢调控, 同源重组, 昆虫病原真菌

Abstract:

The fungal gene fluG is involved in the regulation of conidial generation. However, the role of fluG in the entomopathogenic fungus Metarhizium anisopliae is rarely reported. To investigate the role of fluG in M. anisopliae, we constructed a mutant strain of M. anisopliae with fluG knocked out by DNA homologous recombination to analyze its effect on the sporulation characteristics. The target vector pDHt/sk-fluG-Ben was constructed by using benomyl resistance gene ben as selective marker and sequence of fluG gene flanks as homologous arms. The target vector was transformed into the protoplasts of M. anisopliae by PEG mediating, and the resistant transformants of benomyl were obtained. According to the test of target genes and marker genes, the mutant strains of M. anisopliae with fluG knocked out were determined. Phenotypic analysis showed that the fluG mutant still maintained benomyl resistance after 5 generations of subculture. The mutant colonies looked loose and flocculent, grew significantly slower than the wild type, and could not or only produce a very small number of conidia. These results indicate that the knockout of fluG gene affect the development of the strain and prevent conidia formation, showing fluG an important gene in the regulation of spore-producing of M. anisopliae. This study establishes the basis for further elaborating the regulation mechanism of M. anisopliae conidiation.

Key words: gene knockout, fluG gene, sporulation regulation, homologous recombination, entomopathogenic fungus