PAG-V: W2 - RFLP ANALYSIS OF STRESS RESPONSIVE GENES IN BARLEY

Plant & Animal Genome V Conference

Town & Country Hotel, San Diego, CA, January 12-16, 1997.

W2

RFLP ANALYSIS OF STRESS RESPONSIVE GENES IN BARLEY

MARMIROLI, NELSON, Mariolina Gulli, Aliosha Malcevschi, Elena Maestri
Laboratory of Environmental Biotechnologies, Department of Environmental Sciences, University of Parma, Viale delle Scienze, 43100 Parma, Italy

Stress-responsive-genes (SRG) from wheat and barley, induced in response to heat-shock and dehydration, have been chosen as probes for restriction fragment length polymorphism (SRG-RFLP) analysis of twenty-seven Hordeum vulgare cultivars, encompassing six- and two-rowed, spring and winter genotypes. The SRG chosen for this analysis were: HvHSP17 and TaHSP16.9 encoding for low molecular weight heat shock proteins (LMW-HSP) of barley and wheat, ABA3 encoding for a 20.7-kDa dehydrin of barley, ABA8 encoding for an aldose reductase homologue of barley, and ABA7 encoding for a 18.9-kDa drought-induced protein of unknown function of barley. With these probes a number of polymorphic patterns and alleles were evidenced after Southern hybridization of genomic DNA digested with five different restriction enzymes (RE). Probes derived from heat-shock genes, which are members of a multigenic family, recognized the highest number of bands. Indexes of genetic and phenotypic diversity computed within and between groups of cultivars (two-rowed and six-rowed, winter and spring) have allowed for the determination of high values of genetic differentiation (G_ST). Genetic similarities analyzed by means of principal components analysis, clustering method (UPGMA) and parsimony analysis allowed correct grouping of related cultivars, with few exceptions due to the pedigree of particular cultivars. Starting with the primary sequences of the same SRGs we have elaborated a SRG-AP-PCR (stress responsive genes arbitrarily primed PCR) strategy. These primers allowed the amplification of polymorphic fragments deriving from conserved and/or variable regions of the genes. The SRG-AP-PCR resulted in another way of measuring the variation occurring into these important gene regions, since these regions have a deep physiological and evolutionary meaning.