A number of 10 base primers were screened to identify RAPD polymorphisms among a population of semi wild apricot genotypes that had been collected by Maxine Thompson in 1988. 30 families collected from trees at 6 locations were analyzed. DNA from leaf tissue of 180 plants, ca. 6 genotypes per family, were isolated and tested against 20 primers. Seven primers were identified that produced consistent results with relatively few (thus, scoreable) and consistent bands. DNA was isolated using the cTAB method and the effects of additional CsCl centrifugation isolation were tested. No differences were found. Reaction conditions were tested to ensure consistent results. Considerable RAPD polymorphism was observed in this population. Parsimony analysis is being conducted to assess the relative variation among and within populations and to determine whether collection location had a more significant effect on DNA variation than other factors such as outcrossing or level of heterogeneity within populations.
Y. Gogorcena and D.E. Parfitt
Y. Gogorcena, S. Arulsekar, and D.E. Parfitt
The work reported here is an extension of studies reported in 1990. The general objective was to develop molecular markers for genotype `fingerprinting', with specific reference to possible clonal differences among `Pinot noir' clones. Leaf DNA from 8 cultivars and 9 `Pinot noir' clones were isolated. RFLP and RAPD markers were identified and used to characterize the genotypes. 65 32-P labelled cloned probes were constructed with the pUC18 plasmid and Hind-III digested `Pinot noir' DNA. The probes were tested for their ability to discriminate among the 8 cultivars. 3 probes pGAD10, pGAD15, and pGAD44 showed polymorphisms among the cultivars. pGAD15 was most useful, with 5 polymorphisms for the 8 cultivars. RAPD makers were also tested for `fingerprinting'. Several primers were tested and polymorphisms were identified among cultivars. However, significant problems with repeatability for some bands were observed. Therefore, a series of experiments were conducted to test the effect of season and extraction method. These factors did not account for the inconsistancy which seemed to be more a function of the primer used. None of these studies showed clear evidence that the `Pinot noir' clones tested were geetically different.
Y. Gogorcena, S. Arulsekar, A. Dandekar, and D.E. Parfitt
DNA from 9 cultivars and 5 `Pinot noir' clones were isolated with either the Delaporta or cTAB methods Twenty five 32P label led cloned probes were constructed with the pUC18 plasmid and Hind-III digested `Pinot noir' DNA. Standard methods of isolation and labelling were used. The probes were tested for efficacy of `fingerprinting' the 14 selections. rDNA and cloroplast a/h binding protein probes were also tested. The non-specific probes were not found to be useful as they bound to an excess number of sites and could not be removed from the southern blots, rendering them useless for further analysis. Grape specific probes bound at multiple sites, indicating that multiple fragments were incorporated into the plasmid vectors during library construction. With the greater variability observable with these multi locus probes, significant polymorphism was observed between cultivars, including `Cabernet sauvignon' and `Pinot noir' which were not distinguishable with GPI or PGM isozymes. Variability between clones of `Pinot noir' was observed with several probes, indicating that these selections are different. No variability had been observed at isozyme loci of the `Pinot noir' clones