Disruption of a Magnaporthe grisea cutinase gene.
Using a one-step strategy to disrupt CUT1, a gene for cutinase, cut1- mutants were generated in two strains of Magnaporthe grisea. One strain, pathogenic on weeping lovegrass and barley and containing the arg3-12 mutation, was transformed with a disruption vector in which the Aspergillus nidulans ArgB+ gene was inserted into CUT1. Prototrophic transformants were screened by Southern hybridization, and 3 of 53 tested contained a disrupted CUT1 gene (cut1::ArgB+). A second strain, pathogenic on rice, was transformed with a disruption vector in which a gene for hyg B resistance was inserted into CUT1. Two of the 57 transformants screened by Southern hybridization contained a disrupted CUT1 gene (cut1::Hyg). CUT1 mRNA was not detectable in transformants that contained a disrupted gene. Transformants with a disrupted CUT1 gene failed to produce a cutin-inducible esterase that is normally detected by activity staining on non-denaturing polyacrylamide gels. Enzyme activity, measured either with tritiated cutin or with p-nitrophenyl butyrate as a substrate, was reduced but not eliminated in strains with a disrupted CUT1 gene. The infection efficiency of the cut1- disruption transformants was equal to that of the parent strains on all three host plants. Lesions produced by these mutants had an appearance and a sporulation rate similar to those produced by the parent strains. We conclude that the M. grisea CUT1 gene is not required for pathogenicity.
Sweigard
J A
JA
Central Research and Development, E.I. du Pont de Nemours and Co., Wilmington, DE 19880-0402.
Chumley
F G
FG
Valent
B
B
D001203:Ascomycota / Q000235:genetics*; D015139:Blotting, Southern; D002265:Carboxylic Ester Hydrolases / Q000235:genetics*; D004271:DNA, Fungal / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D005800:Genes, Fungal; D010935:Plant Diseases / Q000382:microbiology; D010957:Plasmids; D012333:RNA, Messenger / Q000235:genetics; D014170:Transformation, Genetic
D004271:DNA, Fungal; D012333:RNA, Messenger; D002265:Carboxylic Ester Hydrolases; C019952:cutinase