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MOLECULAR ANALYSIS OF SEXUAL MORPHOGENESIS IN PYRENOPEZIZA BRASSICAE
MOLECULAR ANALYSIS OF SEXUAL MORPHOGENESIS IN PYRENOPEZIZA BRASSICAE G SINGH and A M. ASHBY Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge, CB2 3EA,UK. Tel: +44 1223 330232, Fax: +44 1223 333953. Background and objectives n and is the cause of Pyrenopeziza brassicae is a hemibiotrophic fungal pathoge light leaf spot, a major disease of winter oilseed rape (Brassica napus subsp. oleifera ). It is heterothallic, having two mating types designated MAT 1-1 and MAT 1-2. Sexual reproduction involves a complex yet co-ordinated pathway of development involving early interaction of the two mating types. The role of sexual development in disease epidemiology is becoming more apparent. We wish to understand the physiological and molecular mechanisms that control sexual morphogenesis in P. brassicae and are currently focusing on three key areas, a) the role of a sex factor (SF) in development, b) the role of the mating type (MAT) loci and c) the influence of SF and the MAT loci on downstream developmentally regulated genes. Materials and methods FPLC, SDS PAGE, DNA extraction, Cloning, PCR and Southern analysis were carried out according to standard protocols Results and conclusions Physiological studies have established the involvement of a potential morphogen, termed sex factor (SF) in controlling development in P. brassicae. SF is a lipoidal component produced during sexual development which has the capacity to promote fruiting body formation and inhibit asexual development in single mating type and crossed cultures. Protein profiles of single mating type isolates induced with SF and individually picked fertile apothecia from a sexual cross revealed the presence of a major protein termed sex factor induced (SFI 1) protein which was absent in asexually reproducing cultures. SFI 1 was purified and internal amino acid sequence used to generate oligonucleotide primers which generated a 700 bp PCR product against P. brassicae MAT 1-2 genomic DNA. This sequence is physically linked to the mating type loci. The MAT loci were cloned following screening of genomic libraries of MAT 1-1 and MAT 1-2 respectively with the SFI 1 probe. The MAT loci are idiomorphic. MAT 1-1 has one open reading frame (ORF) with homology to HMG domain (High Mobility Group DNA binding protein) motifs. MAT 1-2 has three ORFs whose products share homology with alpha -1 proteins (DNA binding proteins), metallothioneins (metal binding proteins) and HMG domain proteins respectively. Current progress in these three areas will be presented. References [1] Ashby AM. 1997. Advances in Botanical Research 24, 31-70.