Plant Regeneration from Callus Cultures of Allium trifoliatum subsp. hirsutum and Assessment of Genetic Stability by Isozyme Polymorphism

Abstract

Plant regeneration from callus cultures of Allium trifoliatum subsp. hirsutum fertile accession F-370, was studied as a means for clonal multiplication and germplasm storage of Allium spp. Callus was induced on in vitro-cultured basal leaf explants. Best proliferation was obtained on modified BDS medium supplemented with (mg/l): 0.75 picloram, 2.0 benzyl adenine, and 900 casein hydrolysate. Shoot and root organogenesis were obtained in 3 to 5 month old subcultured calli, on BDS or MS medium supplemented with (mg/i): either 0.03 picloram or no auxin, 2 BA or 2-isopentenyladenine, and 900 casein hydrolysate. Direct bulb formation, without shoot elongation, occurred on BDS medium with 10 mg/l IBA. Under these conditions, callus formation and organogenesis were not obtained with A. trifoliatum subsp. hirsutum var. sterile, a male-sterile genotype.Most regenerants were phenotypically normal, but some abnormal shoots were also observed, i.e. shoots with vitrified or extremely broad leaves. Isozyme polymorphism analysis of seven proteins in the latter regenerants, and in several callus cultures, revealed significant deviation from the original pattern in esterase, 6-phosphogluconate dehydrogenase and superoxide dismutase. No such deviations were detected in normal regenerated plants.

In vitro propagation and germplasm cold-storage of fertile and male-sterile Allium trifoliatum subsp. hirsutum

“…Electrophoretic patterns of soluble leaf proteins were studied in extracts from field-grown plants and from regenerated plants produced in vitro, following explant cold-storage on several culture media . Isozyme analysis revealed identical banding pattern for all 13 proteins (Viterbo et al ., 1992) in all in vitro-produced plants . This is demonstrated here by the MDH profiles (Fig .…”

“…Growth regulators were added at different concentrations, as detailed in the results . All cultures were kept in a growth room at 24 ± VC, and a 16 h photoperiod of cool-white fluorescent light (9 w/m 2 ) as described previously (Viterbo et al ., 1992) . For standard in vitro propagation, explants were subcultured in a fresh medium every 3 weeks .…”

“…trifoliatum was reported previously (Viterbo et al ., 1992) . In the following, we describe a procedure for in vitro multiplication of fertile and male-sterile genotypes of A .…”

Towards the transformation of onions(Allium cepa)

“…Investigations by van der Valk et al (1992) and Phillips & Hubstenberger (1987) demonstrated that A. fistulosum x A. cepa hybrids were particularly Table 1 Source Allium material and a list of the primary in vitro cultures produced since 1981. Refer to Novak et al (1986), Rabino witch & Brewster (1990), and Peffley (1992) Nair & Seo (1993), Peffley (1992), Pike & Yoo (1990), Rauber & Grunewaldt (1988), Havel & Novak (1988), Phillips & Hubstenberger (1987) Marani et al (1994, Nagakubo et al (1993), Havel & Novak (1988), Nagasawa & Finer (1988a), Ohkoshi et al (1987), Novak (1984), Maggioni & Marchesi (1984), Phillips & Luteyn(1983), Maksoud et al (1983) Buitveld et al (1993, Viterbo et al (1992), Rauber & Grunewaldt (1988), Nagasawa & Finer (1988b), Phillips & Hubstenberger (1987), Phillips & Luteyn (1983), Novak (1983), Oosawaetal. (1981) Seabrook (1994), Kahane et al (1992), Tashiro et al (1985), Phillips & Luteyn (1983) Rabinowitch & Brewster (1990, Havel & Novak (1988), Shahin & Kaneko (1986), Phillips & Luteyn (1983…”

“…The most efficient of the Novak et al (1986), Rabinowitch & Brewster (1990), and Peffley (1992 Keller (1990) Suh& Park (1984 callus systems are probably basal plate callus and embryo/seedling derived callus. Basal plate callus can regenerate de novo (Viterbo et al 1992) and should therefore contain a source of progenitor cells that should be both totipotent and competent to accept DNA. Field-grown onion bulbs are, however, very difficult to surface sterilise which restricts the amount of material readily available for transformation studies.…”