Cloning of the mating type locus from Ascochyta lentis (teleomorph: Didymella lentis) and development of a multiplex PCR mating assay for Ascochyta species.
The mating type (MAT) locus of the lentil pathogen, Ascochyta lentis, was cloned and characterized using thermal asymmetric interlaced and inverse PCR with primers designed to the HMG-box of Ascochyta rabiei. A multiplex PCR assay for mating type was developed based on MAT idiomorph and flanking sequences. Primers were designed to specifically amplify MAT from several Ascochyta spp. including A. pisi, A. fabae and A. viciae-villosae in addition to A. lentis. Four hundred and fifty and 700 bp fragments were amplified from MAT1-1 and MAT1-2 isolates, respectively, and fragment size correlated perfectly with laboratory crosses using mating type tester strains. MAT-specific PCR allowed rapid scoring of mating type in crude DNA extracts from geographically diverse population samples of A. viciae-villosae from California and Washington State, USA. This co-dominant MAT-specific PCR assay will be a valuable tool for studying the population structure, biology and epidemiology of these fungi.
Chérif
Mohamed
M
Laboratoire de Phytopathologie, Institut National Agronomique de Tunisie, Cité Mahrajéne, Tunis, Tunisia.
Chilvers
Martin I
MI
Akamatsu
Hajime
H
Peever
Tobin L
TL
Kaiser
Walter J
WJ
D000595:Amino Acid Sequence; D001203:Ascomycota / Q000235:genetics*; D001483:Base Sequence; D002874:Chromosome Mapping; D003001:Cloning, Molecular; D017931:DNA Primers / Q000235:genetics; D004271:DNA, Fungal / Q000235:genetics; D005656:Fungal Proteins / Q000235:genetics; D049770:Genes, Mating Type, Fungal; D024002:HMGB Proteins / Q000235:genetics; D008969:Molecular Sequence Data; D016133:Polymerase Chain Reaction / Q000379:methods*; D017386:Sequence Homology, Amino Acid
D017931:DNA Primers; D004271:DNA, Fungal; D005656:Fungal Proteins; D024002:HMGB Proteins