Restoration of pathogenicity of a penetration-deficient mutant of Collectotrichum lagenarium by DNA complementation.
Infection by Colletotrichum lagenarium requires formation of an appressorium and of a penetration peg. A mutant, 83,348, defective in morphogenesis of the penetration peg was unable to penetrate into cellulose membranes or infect cucumber leaves. DNA transformation using a wild-type genomic library constructed in pKVB resulted in two transformants, Ppr1 and Ppr2, with restored penetration peg formation, from 2,000 benomyl-resistant transformants. However, penetration into cellulose membranes by these transformants ranged from 30 to 40% compared to greater than 90% by wild-type. Southern-blot hybridization showed that a single copy of a cosmid clone had integrated into the genome of the transformants. A 12.0-kbp fragment of the cosmid vector with the flanking region of wild-type genomic DNA was recovered by plasmid rescue from Ppr1. Using the flanking DNA sequences as a probe for colony blot hybridization, a genomic clone was identified and designated pRP46. Transformants obtained following transformation with pRP46 were able to penetrate cellulose membranes. The penetration frequency of pRP46 transformants ranged from 25 to 65%. Transformants were also pathogenic on cucumber.
Perpetua
N S
NS
Laboratory of Plant Pathology, Faculty of Agriculture, Kyoto University, Japan.
Kubo
Y
Y
Okuno
T
T
Furusawa
I
I
D003001:Cloning, Molecular; D004271:DNA, Fungal / Q000235:genetics*; D005800:Genes, Fungal; D005816:Genetic Complementation Test; D015698:Genomic Library; D003904:Mitosporic Fungi / Q000235:genetics* / Q000472:pathogenicity*; D009154:Mutation; D009693:Nucleic Acid Hybridization; D015183:Restriction Mapping; D014170:Transformation, Genetic; D014675:Vegetables / Q000235:genetics; D014774:Virulence / Q000235:genetics
D004271:DNA, Fungal