Adventitious shoot regeneration was observed using leaf-petiole explants from shoot-proliferating cultures of 'Comet' red raspberry (Rubus idaeus L.). A maximum regeneration rate of 70% (3.7 shoots/explant) was obtained using 4.5-9.1/xM (1-2mg1-1) N-phenyl-N'-l,2,3-thiadiazol-5-ylurea (thidiazuron or TDZ) with 2.5-4.9/xM (0.5-1mgl 1) 1H-indole-3-butanoic acid (IBA) or 2.3 IxM (0.5 mg 1-1) TDZ with 4.9/xM (1 mg 1-1) IBA in modified Murashige-Skoog medium. TDZ was more effective than N-(phenylmethyl)-lH-purin-6-amine (BA) at promoting regeneration in combinations tested with IBA (maximum 50% regeneration rate; 1.8 shoots/explant). Variation in the agar concentration or incubation temperature, orientation or scoring of the leaf-petiole explants and use of separate leaf or petiole explants had no effect on shoot regeneration. Incubation in the dark for 1, 2 or 3 weeks prior to growth in the light did not influence the percent regeneration rate but depressed the number of adventitious shoots. Explant source, from micropropagated shoots or greenhouse-grown plants, had an effect on shoot regeneration that was genotype dependent. Only 8 of 22 (36%) raspberry cultivars were capable of regeneration from leaf explants derived from greenhouse-grown plants.
Isoenzyme staining of horizontal starch gels was used to characterize 23 cultivars and three advanced selections of red raspberry (Rubus idaeus L.). The genotypes were separable using the enzymes malate dehydrogenase, phosphoglucoisomerase, phosphoglucomutase, and triose phosphate isomerase. In addition, staining for isocitrate dehydrogenase and shikimate dehydrogenase revealed polymorphisms in some cultivars. By combining these results with those obtained for 78 previously tested cultivars, 75 of the 104 (72%) genotypes tested were uniquely characterized using the six isoenzymes.
Isoenzyme staining was used to characterize 55 of 78 raspberry cultivars (Rubus idaeus L., R. × neglectus Peck, and R. occidentalis L.). Six enzymes were needed to achieve this characterization: isocitrate dehydrogenase, malate dehydrogenase, phosphoglucoisomerase, phosphoglucomutase, shikimic acid dehydrogenase, arid triose phosphate isomerase. The 23 cultivars that were not uniquely characterized were grouped into eight groups of two and two groups of three and four. Two of these groups comprised black raspberry cultivars, all of which were similar isozymically. Isoenzymes could not distinguish between the cultivar Willamette and a spine-free mutant of the cultivar. Analysis of cultivars obtained from several sources revealed that raspberry cultivar mislabeling exists but is not very prevalent. Regular isoenzyme analysis of raspberry cultivars held by germplasm repositories, certified and other propagators, and breeders is both feasible and advisable for early detection of cultivar mislabeling.
The inheritance of five isoenzymes was studied in red and purple raspberry F1 progenies (Rubus idaeus L. and Rubus × neglectus Peck). Isocitrate dehydrogenase (IDH; EC 1.1.1.42) was a dimeric enzyme present in the cytosol and coded for by one locus (Idh-1). Three of the four crosses analyzed at this locus had deviations from expected ratios, possibly caused by its linkage to a recessive lethal gene. Malate dehydrogenase (MDH; EC 1.1.1.37), phosphoglucoisomerase (PGI; EC 5.3.1.9), and triose phosphate isomerase (TPI; EC 5.3.1.1) were dimeric enzymes with two loci. Each of these three enzymes was present in an organelle and in the cytosol for locus 1 and 2, respectively. Phosphoglucomutase (PGM; EC 2.7.5.1) was monomeric with two loci, Pgm-1 and Pgm-2, located in an organelle and the cytosol, respectively. Each allele at Pgm-1 resulted in the formation of two bands. Although most progenies analyzed supported Mendelian inheritance of polymorphic loci (except for Idh-1), there was a higher than expected number of aberrant segregation ratios observed (18.4%). Analysis of 85 pairs of jointly segregating loci revealed a possible linkage group consisting of Mdh-2, Tpi-2, and Pgm-1.
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