Deletion of the cpku80 gene in the chestnut blight fungus, Cryphonectria parasitica, enhances gene disruption efficiency.
The chestnut blight fungus, Cryphonectria parasitica, and associated virulence-attenuating hypoviruses have emerged as an important model system for studying molecular mechanisms underlying fungal-plant pathogenic interactions. As more gene sequence information becomes available as a result of C. parasitica express sequence tags (ESTs) and ongoing whole genome sequencing projects, the development of an efficient gene disruption system has become an urgent need for functional genomics studies of this important forestry pathogen. Here, we report the cloning of the C. parasitica gene cpku80 that encodes a key component of the nonhomologous end joining DNA repair pathway and the construction of a corresponding deletion mutant strain. The cpku80 mutant was indistinguishable from the parental wild-type strain EP155 in colony morphology, ability to support hypovirus replication, conidiation and virulence. As predicted, the Deltacpku80 strain did exhibit an increased sensitivity to the mutagen methyl methanesulfonate. A test with three selected genes resulted in a gene disruption efficiency of about 80% for the Deltacpku80 strain, a significant increase over the 2-5% levels of homologous recombination generally observed for the wild-type strain EP155. This efficient homologous recombination system provides a powerful tool for large-scale analysis of gene functions in C. parasitica.
Lan
Xiuwan
X
Guangxi Key Laboratory of Subtropical Bioresource Conservation and Utilization, College of Life Science and Technology, Guangxi University, 100 Daxue Road, Nanning Guangxi, 530004, People's Republic of China.
Yao
Ziting
Z
Zhou
Yan
Y
Shang
Jinjie
J
Lin
Haiyan
H
Nuss
Donald L
DL
Chen
Baoshan
B
D001203:Ascomycota / Q000235:genetics* / Q000472:pathogenicity; D003001:Cloning, Molecular; D004260:DNA Repair / Q000235:genetics; D004271:DNA, Fungal / Q000737:chemistry / Q000235:genetics*; D005656:Fungal Proteins / Q000235:genetics / Q000502:physiology*; D015966:Gene Expression Regulation, Fungal; D005800:Genes, Fungal / Q000235:genetics*; D008969:Molecular Sequence Data; D010935:Plant Diseases / Q000382:microbiology*; D017422:Sequence Analysis, DNA; D014774:Virulence / Q000235:genetics
D004271:DNA, Fungal; D005656:Fungal Proteins