Development of Embryoid-like Structures from Cucumis melo L. Callus In Vitro

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Cucurbit Genetics Cooperative Report 6:56-57 (article 29) 1983

H. Young, R. M. Skirvin and J. A. Juvick
Department of Horticulture, University of Illinois, Urbana, IL 61801

Members of the Cucurbitaceae have been grown in tissue culture to yield callus cultures (3) and shoot tip cultures (2, 6). The subject has been partially reviewed by Bottino (1). The production of adventitious shoots from Cucurbita pepo L. has been reported by Jelaska (3). In this paper we report the development of numerous shoots from Cucumis melo L.

Seeds of ‘Harper Hybrid’ muskmelon, ‘Oliver’s Pearl Cluster’ honeydew melon and ‘Takii’s Honey’, a Japanese honeydew melon, were surface disinfested with a 10% Clorox bleach solution and 0.1% Triton X-100 surfactant for 15 minutes followed by two 5-minute rinses with sterile distilled water. The Seeds were transferred onto a proliferation media consisting of Murashige and Skoog (MS) high mineral salts (4) supplemented with (mg/liter): 6-benzylaminopurine (2.0), naphthalene acetic acid (0.1), myo-inositol (100) and Staba vitamins (5). Sucrose was added to the medium at 30 g/liter. The medium was adjusted to a pH of 5.7 with KOH and HCl. Difco Bacto agar was added at 6 g/liter, melted and dispensed into 25 x 150 mm culture tubes at 8 ml/tube, then autoclaved for 15 minutes at 15 pounds pressure per square inch. The cultures were grown at about 23°C under cool white fluorescent illumination of about 300 foot candles with a 16-hour day length. In some cases, the seed coat was partially removed to speed germination. ‘Harper Hybrid’ muskmelon shoot tips were gathered from the field and disinfected in 10% Clorox for 10 minutes. All other procedures were identical to those used for seeds.

Six to eight weeks were required for the seeds to germinate when their seed coats were intact. This period was reduced to 9–14 days with partial seed coat removal. Thirty-three days after germination, the healthy cotyledons developed callus and embryoid-like bodies on their margins. These organized differentiating calli were divided into quadrants and subcultured at 4- to 6-week intervals. Contamination rates with shoot tips were very high and only about 1% of the cultures survived. This could be attributed to the hairy buds and field conditions. The surviving shoot formed callus and embryoid-like structures within 30 days. The embryoids of the cultures developed shoots which emerged from the callus-embryoid mass. These structures can be freely sub-cultured and they proliferate rapidly (Table 1). These cultures are being utilized to explore the role of growth regulators in root and shoot differentiation.

Table 1. Growth and development of Cucumis melo in vitro.
No.
explants		 No.
germinating
or surviving		 Number of embryoids/mo.
Explant source 1st
subculture		 Subsequent
subcultures
Seeds
Oliver’s Pearl Cluster		 10 1 2.2 X 4 X
   Takii’s Honey 10 2 2.5 X 4 X
Shoot tips
Harper Hybrid		 85 1 2  X 4 X

Literature Cited

  1. Bottino, P. J. 1981. The tissue culture of vegetable crops. In: Conger, B. V. (ed.), Cloning agricultural plants via in vitro techniques. Chemical Rubber Company Press, Inc., Boca Raton, Florida. pp. 141–164.
  2. Handley, L. W. and Chambliss. 1979. In vitro propagation of Cucumis sativus L. HortScience 14(1):22–23.
  3. Jelaska, S. 1974. Embryogenesis and organogenesis in pumpkin explants. Physiol. Plant. 31:257–261.
  4. Murashige, T. and F. A. Skoog. A revised medium for rapid growth and bioassays with tobacco cultures. Physiol. Plant. 15:473–497.
  5. Staba, J. E. 1969. Plant tissue culture as a technique for the phytochemist. Recent Adv. in Phytochem. 2:80.
  6. Wehner, T. C. and R. D. Locy. 1981. In vitro adventitious shoot and root formation of cultivars and lines of Cucumis sativus L. HortScience 16 (6):759–760.