Genome analysis of the plant pathogenic ascomycete Leptosphaeria maculans; mapping mating type and host specificity loci.

Abstract A genetic and physical map has been developed for the loculoascomycete Leptosphaeria maculans, a pathogen of oilseed Brassicas. The genetic map was constructed from 58 F(1) progeny and comprises 155 amplified fragment length polymorphic (AFLP) markers, three random amplified polymorphic DNA (RAPD) markers, the mating type locus and a host specificity locus conferring the ability to form lesions on Brassica juncea. Twenty-one linkage groups, 5 pairs, and 18 unlinked markers were assigned, and the genome size was 1520 cM. Pulsed field gel electrophoresis experiments showed that the parental isolates each had 16 chromosomes and a genome size of about 33.5 Mb. Attempts to anchor a large number of markers to chromosomes were hampered by difficulties in converting AFLPs into RFLP markers, and because many markers bound to every chromosome, indicating that L. maculans has a high level of dispersed repetitive sequences. This fungus displays chromosomal length polymorphisms, but in the cross examined, the linkage and physical maps were essentially congruent and there was no evidence of translocations. The host specificity locus is 18 cM from the nearest AFLP marker and is located on a chromosome sized 1.85 Mb in the virulent parent. The mating type locus is on a chromosome sized 2.6 Mb and coincident on an AFLP marker amplified from the virulent parent. The derived amino acid sequence of part of this marker has some conserved amino acids present in the High Mobility Group DNA binding domain of MAT-2 mating type genes of other ascomycetes.

Organisation of ribosomal DNA in the ascomycete Leptosphaeria maculans.

In the ascomycete Leptosphaeria maculans tandem repeats of ribosomal DNA (rDNA) are restricted to one or two particular chromosomes of the 15 chromosomes of 19 field isolates examined. Ribosomal DNA can account for size differences of 35% between homologous chromosomes in a particular tetrad. During crossing, no detectable recombination between blocks of tandem repeats, nor changes in their size occur. The organisation of rDNA in L. maculans differs from many other haploid fungi. Firstly, sequence heterogeneity occurs within tandem repeats of rDNA; regularly spaced Sal 1 sites (0.25 Mb apart) are present within a 1.4 Mb block of tandem repeats. Secondly, individual isolates have different-sized rDNA repeats; this variation occurs in the non-transcribed intergenic spacer region. Thirdly, there is a wide range in the copy number of the rDNA repeat (from 56 to 225) amongst only four field isolates examined.