FORWARD AND REVERSE GENETIC ANALYSIS IN MEDICAGO TRUNCATULA

In order to set up a functional genomics tool for forage legumes about 1000 Medicago truncatula lines were mutagenized with the tobacco Tnt1 retroelement. As a reverse genetic approach about 100 flanking regions from 13 random lines were isolated and sequenced. About 40% of sequences showed similarity with known genes. Phenotypic screenings were carried out for several traits including plant architecture and forage quality. Three lines, B21, B1 and C43, segregating the mutant phenotype in a ratio compatible with a single locus inheritance, were chosen for further molecular characterization. Tnt1 flanking regions were isolated and sequence information was used to follow insertion sites segregation. Here we report the preliminary characterization of the B21 mutant line which shows a strikingly dwarf plant with small leaves and short internodes. Southern analysis showed that parental B21 lines contains at least 5 insertions. The host flanking region of 4 insertion sites were cloned by Inverse PCR and S-SAP and further caharcaterized. Homologies were found to: HCR4, Peptidyl-prolyl cis-trans isomerase, CHS and the Mth2-81g19 sequenced BAC clone of Medicago truncatula. As far none of the characterized insertions co-segregated with the mutant phenotype, therefore isolation of more flanking regions is in progress.