The nuclear rDNA intergenic spacer of the ectomycorrhizal basidiomycete Laccaria bicolor: structural analysis and allelic polymorphism

The nuclear rDNA intergenic spacer (ICS) of the ectomycorrhizal basidiomyce te Laccaria bicolor was amplified and sequenced to identify the source of i ts intraspecific polymorphism The ICS was amplified by PCR in several L. bi color strains and shown to exhibit multiple bands and length polymorphism. The IGS loci were shown to segregate in a 1:1 ratio within haploid progenie s in three dikaryotic strains, suggesting that divergent ICS haplotypes wer e present in the two nuclei of these strains. The two haplotypes of L. bico lor S238N were sequenced: the beta-haplotype was 4160 bp in length, whereas the size of the alpha-haplotype was estimated to be about 4700 bp. These r epresent the largest published fungal ICS sequences to date. These sequence s can be subdivided into three main regions, IGS(1), 55 rDNA and IGS(2). Th e ICS sequences are AT-rich and contain numerous occurrences of three types of subrepeats (e.g. T(2)AC(3)). The length polymorphism, observed between the ICS sequence of the alpha- and beta-haplotypes, results from the insert ion of various numbers of a 71 bp subrepeat, called B, occurring in IGS(2). This variation in subrepeat number suggests that the two haplotypes result ed from unequal cross-overs. The L. bicolor IGS was aligned with ICS sequen ces of two other Tricholomataceae (i.e. Tricholoma matsutake and Collybia f usipes). No sequence similarity was observed between these IGSs, but homolo gous subrepeats were found in L. bicolor and T. matsutake. Analysis of ICS length polymorphism is therefore an efficient tool for investigating geneti c relationships between genets and within progenies in natural fungal popul ations.