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Cucurbit Genetics Cooperative Report 4:20-22 (article 10) 1981

Tissue Culture Propagation of Field-Grown Cucumber Selections

T. C. Wehner and R. D. Locy

North Carolina State University, Raleigh, NC 27650

One potential application of tissue culture techniques to cucumber breeding is in the propagation of selections made in the field. This would replace vegetative propagation by rooted cuttings (4). Callus has been induced to form from all vegetative and reproductive parts of the cucumber plant: root, stem, leaf blade, petiole, tendril, flower parts, and seedling hypocotyls and cotyledons (1). Callus has also been produced from leaf protoplasts (2).

One method of propagation would involve producing callus from the selected plants, multiplying the callus, and then regenerating plants from the callus. The best source of tissue for use in propagating selections is leaf tissue, since it is available throughout the life of the plant, is available in large quantities, and, if taken near the growing point, represents younger and more rapidly growing tissue than most other plant parts. The major problem in vegetatively propagating cucumber selections from the field is eliminating external and internal contamination without killing the tissue. Our objective was to evaluate a tissue culture approach to propagation of genetically diverse cucumbers grown in the field under normal summer disease load. In addition, we evaluated sodium hypochlorite (chlorine bleach) for use in sterilizing tissues to be cultured in vitro.

Thirty diverse Cucumis plant introduction (PI) lines were planted at Raleigh, NC on June 27. All but two of the PI lines used were Cucumis sativus accessions (Table 1). The exceptions are PI 299570 which is C. africanus, and PI 292190 which is C. metuliferus. After 41 days, the youngest fully-opened leaf was collected from 3 plants of each PI line. Leaves were then sterilized in a 17% solution of laundry bleach (1 bleach:5 water) for 15 to 42 min (depending upon the sterilizing treatment), and then 3 mm square pieces of blade or petiole tissue were cut and placed on the tissue culture medium with 5 tissue pieces per petri plate. The medium contained Murashige-Skoog (3) salts and vitamins with 0.1 mg/l naphthalene acetic acid, 1 mg/l N6-benzylamino purine, and 3% sucrose. A completely random experimental design was used with 30 lines, 2 tissues (leaf blade and leaf petiole), and 2 replications (petri plates). The plates were evaluated for contamination and callus production after 34 days in culture.

Tissue from leaf blades produced callus with a higher rate of success than that from leaf petioles (Table 2). Uncontaminated callus was obtained from 25 of the 30 lines used, with the loss of callus being due to either internal contamination of the tissue or to killing of the tissue for callus during sterilization. Tissue from leaf petioles is not an efficient source of tissue for callus production because of its high rate of internal contamination (Table 3), and its low rate of clean callus production (Table 2). Leaf blades should be sterilized in 17% laundry bleach for about 20 min to eliminate as much contamination as possible without killing the tissue (Table 3). Better than 60% of the petiole and blade pieces of the C. africanus and C. metuliferus PI lines produced clean callus.

It appears that, with further refinements, the system outlined here could be used to reliably establish callus cultures of selections made from field-grown cucumber plants.

Table 1. Plant introduction (PI) lines, and their origins, used in this study.

PI

Origin

PI

Origin

PI

Origin

92806

China

222099

Afghanistan

299570z

South Africa

165499

India

223841

Philippines

342950

Denmark

167050

Turkey

224668

Korea

344442

Iran

177363

Syria

228808

Canada

355052

Israel

181756

Lebanon

249561

Thailand

356809

U. S. S. R.

183224

Egypt

257286

Spain

357837

Yugoslavia

205995

Sweden

264226

France

358813

Malaysia

206043

Puerto Rico

267745

Brazil

360939

Netherlands

212059

Greece

283899

Czechoslovakia

369717

Poland

212233

Japan

288990

Hungary

292190y

Transvaal

z Cucumis africanus
y Cucumis metuliferus

Table 2. Callus production from leaf tissue collected from 30 field-grown Cucumis lines.z

Tissue source

Clean callus (%)

Contaminated callus (%)

Dead tissue (%)

Petiole

38

32

30

Blade

58

7

35

z Youngest fully-opened leaf harvested from 41-day-old plants growing at Raleigh, NC.

Table 3. Percent loss of leaf tissue pieces to contamination or to killing by sterilization.

 

Tissue source z

 

Leaf petiole

Leaf blade

Time in sodium hypochlorite (min.)

Dead

Contaminated

Dead

Contaminated

15

0

77

13

33

18

10

33

7

7

21

47

20

0

7

24

30

67

33

0

27

43

10

47

0

30

47

17

40

0

33

30

17

13

0

36

43

23

73

7

39

23

17

53

7

42

33

37

60

13

z Youngest fully-opened leaf harvested from 41-day-old plants growing at Raleigh, NC.

Literature Cited

  1. Alsop, W. R., W. W. Cure, G. F. Evans, and R. L. Mott. 1978. Preliminary report on in vitro propagation of cucumber. Cucurbit Genetics Coop. Rpt. 1:1-2.
  2. Coutts, R. H. A. and K. R. Wood. 1977. Improved isolation and culture methods for cucumber mesophyll protoplasts. Plt. Sci. Lett. 9:45-57.
  3. Murashige, T. and F. Skoog. 1962. A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiol. Plant. 15:473-497.
  4. Shehatta, M. A., D. W. Davis, and P. E. Reed. 1974. Vegetative propagation of cucumber. HortScience. 9:575-576.
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Page citation: Wehner, T.C., Cucurbit Genetics Cooperative;
Created by T.C. Wehner and T. Ng, 1 June 2005; design by C.T. Glenn;
send questions to T.C. Wehner; last revised on 23 October, 2009