Methylation-sensitive RFLP analysis of the apospory-specific genomic region (ASGR) of Paspalum notatum

RODRIGUEZ MP; QUARIN CL; ORTIZ JPA

Buenos Aires

Simposio; The 6th International Symposium of the molecular breeding of forage and turf; 2010

Molecular Breeding of Forage and Turf International Committee

The addition of a methyl group to the C5 position of a cytosine residue is the most common DNA modification in plants and animals. It is involved in numerous biological processes, including embryogenesis and development, genomic imprinting, silencing of transposable elements and regulation of gene transcription. Paspalum notatum Flugge is a polymorphic species with sexual diploids and apomictic triploids, tetraploids and pentaploids. The diploid form (2n=2x=20) Paspalum notatum var. saurae Parodi (Pensacola bahiagrass) is native to a relatively small area of Central Argentina. It reproduces sexually and is highly self-incompatible. Tetraploid races (2n=4x=40), usually considered to be the typical form of the species in botanical terms, constitute the most common cytotype. In a recent work we developed a genetic linkage map of the species based on AFLP and RFLP molecular markers where the linkage group carrying the apospory locus (apo-locus) was identified. A single RFLP (rice probe C 1069), 2 RAPD and 20 AFLP markers, co-segregating strictly with the trait, defined the apospory specific genomic region of the species. Methylation-sensitive genomic hybridization with clones M3 (an AFLP-derived clone) and C1069 were carried out on DNA samples from the apomictic genotype Q4117 and the sexual one Q4188 digested with both HpaII and MspI isoschizomers. To identify putative polymorphism associated with apospory, bulked samples of 10 aposporous and 10 non-aposporous F1 plants derived from them were included. Hybridization results showed polymorphisms between the isoschizomers in all samples tested, indicating that the region is methylated in both sexual and apomictic plants. Furthermore, the methylation analysis of Fl plants with probe C1069 showed polymorphisms associated with the reproduction mode. Although C1069 is a portion of a conserved coding region of a transposase enzyme, a functional component of transposable elements known to be inactivated in plants to prevent their mobilization, we observed at least partial demethylation of this locus on the aposporous plants.