Cucurbit Genetics Cooperative Report 1:1-2 (article 1) 1978
Preliminary Report on In Vitro Propagation of Cucumber
W. R. Alsop, W. W. Cure, G. F. Evans, and R. L. Mott
North Carolina State University, Raleigh, NC 27650
Preliminary investigations on the feasibility of in vitro propagation of cucumber (Cucumis sativus L.) have been conducted in this laboratory. Shoot regeneration from callus has not been reported for this species although some success has been reported with related species (1, 2). Maciejewska-Potapczykowa et al. (3) have observed callus growth as well as in vitro axillary and floral bud growth in cucumber. However, their studies emphasized possible mechanisms of sex determination rather than propagation. Coutts and Wood have also tried unsuccessfully to get shoots from callus (4). We have conducted experiments to determine an appropriate medium for callus growth and organogenesis, and to investigate the possibilities of in vitro propagation using organ explants.
Explants obtained from 6-7 week old greenhouse-grown cucumber plants were surface sterilized in 10% Clorox solution with a drop of liquid detergent added followed by water rinses. Media including Murashige and Skoog (5), Greshoff and Doy (6), and White's (7) have been employed with various combinations of coconut milk, casein hydrolysate, and growth regulators. The best callus growth was observed on Murashige and Skoog medium supplemented with 10% coconut milk (v/v) and naphthalene acetic acid (NAA) (0.1 mg/l) on 1% agar solidified media at 21°C under approximately 2,000 lux fluorescent light. Callus has been initiated from stems, roots, tendrils, petioles, leaves, and flowers, as well as seedling hypocotyls and cotyledons. On this medium, callus frequently gives rise to adventitious roots, which then grow rapidly.
To date, our attempts to induce shoot formation from callus have been unsuccessful. Single and combined additions of NAA and benzylaminopurine (BAP) in concentrations of O to 5 and 0 to 10 mg/l respectively have been tried. The addition of coconut milk was beneficial regardless of other media components. BAP became toxic at concentrations of 10 mg/l or higher, and most vigorous callus growth was observed in the absence of BAP. Callus grown on NAA (0.1 mg/l) and BAP (0-1 mg/l) exhibited a tendency to organize into round knobs with green centers. However, no further organization has been achieved.
A propagation scheme which may be of use to breeder utilizes the induction of axillary bud development from the nodes of excised stem segments. The crevices at the leaf axil make surface sterilization of excised nodes particularly difficult, especially from field-grown plants. Uncontaminated node explants cultured on White's salts with 30.0 g/l sucrose and 300 mg/l casein hydrolysate typically formed callus from the pith of the basal portion of the stem, and adventitious roots were observed in just 3 or 4 days. Vegetative axillary buds grew out after approximately 2 weeks. The plants grew to be 6 to 8 nodes in length and flowered under continuous fluorescent light.
We are continuing experiments aimed toward shoot regeneration from callus. However, from a practical point of view and from these preliminary experiments, it seems that a workable in vitro propagation system for clonal propagation of Cucumis sativus L. could make use of the many axillary buds which already exist at the nodes of the cucumber plant. Where desired, further increase could be obtained by successively subculturing nodes excised from the already sterile shoots formed in vitro.
- Jelaska, Sibila. 1972. Embryoid formation by fragments of cotyledons and hypocotyls in Cucurbita pepo. Planta 103:278-280.
- Jelaska, Sibila. 1974. Embryogenesis and organogenesis in pumpkin explants. Physiol. Plant. 31:257-261.
- Maciejewska-Potapczykowa, W., A. Rennert and E. Milewska. 1972. Callus induction and growth of tissue cultures derived from cucumber plant organs of four different sex types. Acta. Societatis Botanicorum Polloniae 41:329-339.
- Coutts, R. H. A. and K. R. Wood. 1977. Improved isolation and culture methods for cucumber mesophyll protoplasts. Plt. Sci. Lett. 9:45-57.
- Murashige, T. and F. Skoog. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15:473-497.
- Sommer, H. F., C. L. Brown and P. P. Kormanik. 1975. Differentiation of plantlets in longleaf pine (Pinus palustris Mill.) tissue cultures in vitro. Bot. Gaz. 136:196-200.
- White, P. R. 1954. The cultivation of plant and animal cells. Ronald Press, New York.