Although methylation levels are quite low in fungi, conspicuous methylation has been reported in several systems: the best examples are found in Neurospora crassa associated with duplicated sequences (repeat-induced point mutation: RIP), in genomic repeated sequences, mainly rDNA, and in some quiescent stages. In order to study DNA methylation in Cryphonectria parasitica, Botrytis cinerea and Pyrenophora graminea, their total genomic DNA has been analysed both by restriction analysis, based on methylation dependent isoschizomers (HpaII and MspI) and by high pressure liquid chromatography (HPLC). DNA methylation of specific genomic sites has been evaluated in Pyrenophora graminea IGS region; preliminary results based on restriction and Southern blot analysis suggest further studies should be carried out with a wider range of restriction enzymes. The possible involvement of methylation in regulation of gene expression in fungi is evaluated by looking for methylated sites in active or inactive promoters of cloned genes for an ABC transporter in Botrytis cinerea and for laccase in Cryphonectria parasitica. Finally a search will be undertaken in the same fungi to identify genes coding for methyltransferases, enzymes involved in transfer of methyl groups to specific cytosine residues in DNA.
Search for DNA methylation in Cryphonectria parasitica, Botrytis cinerea and Pyrenophora graminea
VERGARA, Mariarosaria;
2000
Abstract
Although methylation levels are quite low in fungi, conspicuous methylation has been reported in several systems: the best examples are found in Neurospora crassa associated with duplicated sequences (repeat-induced point mutation: RIP), in genomic repeated sequences, mainly rDNA, and in some quiescent stages. In order to study DNA methylation in Cryphonectria parasitica, Botrytis cinerea and Pyrenophora graminea, their total genomic DNA has been analysed both by restriction analysis, based on methylation dependent isoschizomers (HpaII and MspI) and by high pressure liquid chromatography (HPLC). DNA methylation of specific genomic sites has been evaluated in Pyrenophora graminea IGS region; preliminary results based on restriction and Southern blot analysis suggest further studies should be carried out with a wider range of restriction enzymes. The possible involvement of methylation in regulation of gene expression in fungi is evaluated by looking for methylated sites in active or inactive promoters of cloned genes for an ABC transporter in Botrytis cinerea and for laccase in Cryphonectria parasitica. Finally a search will be undertaken in the same fungi to identify genes coding for methyltransferases, enzymes involved in transfer of methyl groups to specific cytosine residues in DNA.I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
