Medium effects on micropropagation and genetic stability of Citrullus lanatus oleaginous type

DOI: 10.4236/as.2013.47A005   PDF   HTML     3,821 Downloads   6,027 Views   Citations


To regenerate adventitious shoots from the cotyledon proximal parts of Citrullus lanatus (Thunb.) Matsum. and Nakai ssp. mucosospermus (Fursa) oleaginous type, different concentrations of MS mineral elements, sucrose, 6-benzylaminopurine (BAP) and agar were tested. Shoot induction proved to depend on the interaction between levels of sucrose, BAP and MS mineral elements in the medium. The medium containing 3/2 strength of MS mineral elements, 35 g/l sucrose and 1 mg/l BAP solidified with 6 g/l agar allowed the production of numerous shoots without a callus phase. After 3 weeks of culture, 76.7% of the cotyledon proximal parts induced shoots with an average of 12.26 shoots per explant and a mean shoot length of 17.13 mm. The induced shoots were directly rooted and thus complete plants ready for acclimatization were obtained using a two steps procedure. Depending on the genotype, the shoot induction from cotyledon proximal parts ranged from 54% to 96%. Rooted plantlets were acclimatized and transferred to field, where they grew well, developed flowers and fruits like seeded plants. The assessment of the genetic stability of the in-vitro-regenerated plantlets by means of an Amplified Fragment Length Polymorphism (AFLP) analysis with the combination of 5 primers revealed no differences between regenerated plantlets and mother plants.

Share and Cite:

Gnamien, Y. , Zoro Bi, I. , Kouadio, Y. , Brostaux, Y. and Baudoin, J. (2013) Medium effects on micropropagation and genetic stability of Citrullus lanatus oleaginous type. Agricultural Sciences, 4, 32-44. doi: 10.4236/as.2013.47A005.

Conflicts of Interest

The authors declare no conflicts of interest.


[1] Gusmini, G., Wehner, T.C. and Jarret, R.L. (2004) Inheritance of Egusi seed type in watermelon. Journal of Heredity, 95, 268-270. doi:10.1093/jhered/esh031
[2] Gnamien, Y.G., Zoro Bi, I.A., Djè, Y., Toussaint, A. and Baudoin, J.P. (2010) Determination of a suitable protocol for indigenous oilseed cucurbits plant regeneration. Tropicultura, 28, 217-225.
[3] Kortse, P.A., Oladiran, J.A. and Msaakpa, T.S. (2012) Effect of season and fruit size on the quality of “egusi” melon [Citrullus lanatus (Thunb) Matsum and Nakai] seed. ARPN Journal of Agricultural and Biological Science, 7, 110-116.
[4] Zoro Bi, I.A., Koffi, K.K. and Djè, Y. (2003) Caractérisation botanique et agronomique de trois espèces de cucurbites consommées en sauce en Afrique de l’Ouest: Citrullus sp., Cucumeropsis mannii Naudin et Lagenaria siceraria (Molina) Standl (Botanical and agronomical characterization of three species of cucurbit consumed as sauce in west Africa: Citrullus sp., Cucumeropsis mannii Naudin and Lagenaria siceraria (Molina) Standl.). Biotechnolgy, Agronomy, Society and Environment, 7, 189-199.
[5] Schippers, R. (1997) Egusi. In: IPGRI, Ed., African Indigenous Vegetables, IPGRI/NRI, IPGRI Workshop Proceeding, Limbe, 93-112.
[6] Zoro Bi, I.A., Koffi, K.K., Djè, Y., Malice, M. and Baudoin, J.P. (2006) Indigenous cucurbits of Cote d’Ivoire. Review of their genetic resources. Science & Nature, 3, 1-9.
[7] Achu, M.B., Fokou, E., Tchiengang, Fotso, M. and Mbiapo, F.T. (2005) Nutritive value of some Cucurbitaceous oilseeds from different regions of Cameroon. African Journal of Biotechnology, 4, 1329-1334.
[8] Loukou, A.L., Gnakri, D., Djè, Y., Kippré, A.V., Malice, M., Baudoin, J.P. and Zoro Bi, I.A. (2007) Macronutrient composition of three cucurbit species cultivated for seed consumption in Cote d’Ivoire. African Journal of Biotechnology, 6, 529-533.
[9] Dalziel, J.M. (1937) The useful plants of west tropical Africa. The Crown Agents for the Colonies, London.
[10] Maggs-Kolling, G.L. and Christiansen, J.L. (2003) Variability in Namibian landraces of watermelon (Citrullus lanatus). Euphytica, 132, 251-258. doi:10.1023/A:1025053331528
[11] Minsart, L.-A., Zoro Bi, I.-A., Djè, Y., Baudoin, J.-P., Jacquemart, A.-L. and Bertin, P. (2011) Set up of simple sequence repeat markers and first investigation of the genetic diversity of West-African watermelon (Citrullus lanatus ssp. vulgaris oleaginous type). Genetic Resources and Crop Evolution, 58, 805-814. doi:10.1007/s10722-010-9617-x
[12] Obiagwu, C.J. and Odiaka, N.I. (1995) Fertilizer schedule for yield of fresh fluted pumpkin (Telfairia occidentalis) grown in lower Benue river basin of Nigeria. Indian Journal of Agricultural Sciences, 65, 98-101.
[13] Ndabalishye, I. (1995) Agriculture vivrière ouest-africaine à travers le cas de la Cote d’Ivoire (West African food crops through the case of Cote d’Ivoire). IDESSA, Bouaké, 383.
[14] Dudu, P., Lale, N.E.S. and Okiwelu, S.N. (1996) Susceptibility of three physical forms of three oilseeds Oryzaephilus mercator and the effects of infestation on seed quality. Postharvest Biology and Technology, 7, 277-283. doi:10.1016/0925-5214(95)00043-7
[15] Djè, Y., Kouonon, L.C., Zoro Bi, I.A., Gnamien, Y.G. and Baudoin, J.P. (2006) étude des caractéristiques botaniques, agronomiques et de la biologie florale du melon africain (Cucumis melo L. var. agrestis Naudin, Cucurbitaceae) (Study of the botanical, agronomic characteristics and the floral biology of the African melon (Cucumis melo L. var. agrestis Naudin, Cucurbitaceae). Biotechnolgy, Agronomy, Society and Environment, 10, 109-119.
[16] Nerson, H. (1991) Fruit age and seed extraction procedures affect germinability of cucurbit seeds. Seed Science Technology, 19, 185-195.
[17] Nerson, H. (2002) Effects of seed maturity, extraction practices and storage duration on germinability in watermelon. Scientia Horticulturae, 93, 245-256. doi:10.1016/S0304-4238(01)00332-6
[18] Pitrat, M. (2008) Melon. In: Prohens, J. and Nuez, F., Eds., Vegetables I: Asteraceae, Brassicaceae, Chenopodicaceae, and Cucurbitaceae. Springer Science+Business Media, LLC, New York, 283-315.
[19] Suratman, F., Huyop, F. and Parveez, G.K.A. (2009) In vitro shoot regeneration of Citrullus vulgaris schrad (watermelon). Biotechnology, 8, 393-404. doi:10.3923/biotech.2009.393.404
[20] Choi, J.Y., Shin, Y.S., Chung, Y.S. and Hyung, N.I. (2012) An efficient selection and regeneration protocol for Agrobacterium-mediated transformation of oriental melon (Cucumis melo L. var. makuwa). Plant Cell, Tissue and Organ Culture, 110, 133-140. doi:10.1007/s11240-012-0137-6
[21] Chuang, S.J., Chen, C.L., Chen, J.J., Chou, W.Y. and Sung, J.M. (2009) Detection of somaclonal variation in micro-propagated Echinacea purpurea using AFLP marker. Scientia Horticulturae, 120, 121-126. doi:10.1016/j.scienta.2008.09.020
[22] De la Puente, R., González, A.I., Ruiz, M.L. and Polanco, C. (2008) Somaclonal variation in rye (Secale cereale L.) analyzed using polymorphic and sequenced AFLP markers. In Vitro Cellular and Developmental Biology—Plant, 44, 419-426. doi:10.1007/s11627-008-9152-z
[23] Murashige, T. and Skoog, F. (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiologia Plantarum, 15, 473-497. doi:10.1111/j.1399-3054.1962.tb08052.x
[24] Box, G. and Behnken, D. (1960) Some new three level designs for the study of quantitative variables. Technometrics, 2, 455-475. doi:10.1080/00401706.1960.10489912
[25] Koffi, K.K., Anzara, G.K., Malice, M., Djè, Y., Bertin, P., Baudoin, J.-P. and Zoro Bi, I.A. (2009) Morphological and allozyme variation in a collection of Lagenaria siceraria (Molina) Standl. from Cote d’Ivoire. Biotechnology, Agronomy, Society and Environment, 13, 257-270.
[26] Lacape, J.M., Dessauw, D., Rajab, M., Noyer, J.L. and Hau, B. (2007) Microsatellite diversity in tetraploid Gossypium germplasm: Assembling a highly informative genotyping set of cotton SSRs. Molecular Breeding, 19, 45-58. doi:10.1007/s11032-006-9042-1
[27] Chalhoub, B.A., Thibault, S., Laucou, V., Rameau, C., Hofte, H. and Cousin, R. (1997) Silver staining and recovery of AFLP TM amplification products on large denaturing polyacrylamide gels. Biotechniques, 22, 216-220.
[28] Dagnelie, P. (1998) Theoretical and applied statistics Volume 2: Statistical inference for one and two dimensions. De Boeck & Larcier, Paris et Bruxelles, 508.
[29] Plader, W., Malepszy, S., Burza, W. and Rusinowski, Z. (1998) The relationship between the regeneration system and genetic variability in the cucumber (Cucumis sativus L.). Euphytica, 103, 9-15. doi:10.1023/A:1018359726626
[30] Zhang, Y., Zhou, J., Wu, T. and Cao, J. (2008) Shoot regeneration and the relationship between organogenic capacity and endogenous hormonal contents in pumpkin. Plant Cell Tissue and Organ Culture, 93, 323-331. doi:10.1007/s11240-008-9380-2
[31] Canli, F.A. and Tian, L. (2008) Regeneration of adventitious shoots from mature stored cotyledons of Japanese plum (Prunus salicina Lind1). Scientia Horticulturae, 120, 64-69. doi:10.1016/j.scienta.2008.09.017
[32] Thomas, T.D. and Sreejesh, K.R. (2004) Callus induction and plant regeneration from cotyledonary explants of ash gourd (Benincasa hispida L.). Scientia Horticulturae, 100, 359-367. doi:10.1016/j.scienta.2003.06.001
[33] Huq, A., Akter, S., Islam, S. and Khan, S. (2012) In vitro micropropagation of pointed gourd (Trichosanthes dioica Roxb.) from shoot tip and nodal segment. Bangladesh Journal of Scientific and Industrial Research, 47, 217-222. doi:10.3329/bjsir.v47i2.11457
[34] Lou, H. and Kako, S. (1994) Somatic embryogenesis and plant regeneration in cucumber. HortScience, 29, 906-909.
[35] Cousineau, J.C. and Donnelly, D.J. (1991) Adventitious shoot regeneration from leaf explants of tissue cultured and greenhouse-grown raspberry. Plant Cell Tissue and Organ Culture, 27, 249-255. doi:10.1007/BF00157588
[36] Ladyman, J.A.R. and Girard, B. (1992) Cucumber somatic embryo development on various gelling agents and carbohydrate sources. HortScience, 27, 164-165.
[37] Amiri, M.E. (2006) Effect of mineral concentration on in vitro explant growth of almond (Prunus amygdalus var. Binazir). Journal of Applied Horticulture, 8, 62-64.
[38] Bejoy, M., Dan, M. and Anish, N.P. (2006) Factors affecting the in vitro multiplication of the endemic zingiber Curcuma haritha mangaly and sabu. Asian Journal of Plant Science, 5, 847-853. doi:10.3923/ajps.2006.847.853
[39] Brar, M.S., Al-Khayri, J.M., Morelock, L.T.E. and Anderson, E.J. (1999) Genotypic response of cowpea Vigna unguiculata (L.) to in vitro regeneration from cotyledon explants. In Vitro Cellular and Developmental Biology-Plant, 35, 8-12. doi:10.1007/s11627-999-0002-4
[40] Vandemoortele, J.L., Billard, J.E., Boucaud, J. and Gaspar, T. (1999) Evidence for an interaction between basal medium and plant growth regulators during adventitious or axillary shoot formation of cauliflower. In Vitro Cellular and Developmental Biology-Plant, 35, 13-17. doi:10.1007/s11627-999-0003-3
[41] George, E.F., Hall, M.A. and De Kler, G.J. (2008) Plant propagation by tissue culture. 3rd Edition, Springer, Dordrecht.
[42] Srivastava, D.K., Andrianov, V.M. and Piruzian, E.S. (1989) Tissue culture and plant regeneration of watermelon. Plant Cell Reports, 8, 300-302. doi:10.1007/BF00274135
[43] Compton, M.E. and Gray, D.J. (1993) Shoot organogenesis and plant regeneration from cotyledons of diploid, triploid, and tetraploid watermelon. Journal of American Society for Horticultural Science, 118, 151-157.
[44] Abrie, A.L. and Van Staden, J. (2001) Development of regeneration protocols for selected cucurbit cultivars. Plant Growth Regulation, 35, 263-267. doi:10.1023/A:1014415419158
[45] Niedz, R.P., Smith, S.S., Dunbar, K.B., Stephens, C.T. and Murakishi, H.H. (1989) Factors influencing shoot regeneration from cotyledonary explants of Cucumis melo. Plant Cell Tissue and Organ Culture, 18, 313-319. doi:10.1007/BF00043400
[46] Ananthakrishnan, G., Xia, X., Elman, C., Singer, S., Paris, H.S., Gal-On, A. and Gaba, V. (2003) Shoot production in squash (Cucurbita pepo) by in vitro organogenesis. Plant Cell Reports, 21, 739-746.
[47] Han, J.S., Oh, D.G., Mok, I.G., Park, H.G. and Kim, C.K. (2004) Efficient plant regeneration from cotyledon explants of bottle gourd (Lagenaria siceraria Standl.). Plant Cell Reports, 23, 291-296. doi:10.1007/s00299-004-0846-3
[48] Krug, M.G.Z., Stipp, L.C.L., Rodriguez, A.P.M. and Mendes, B.M.J. (2005) In vitro organogenesis in watermelon cotyledons. Pesquisa Agropecuária Brasileira, 40, 861-865. doi:10.1590/S0100-204X2005000900004
[49] Compton, M.E., Gray, D.J. and Gaba, V.P. (2004) Use of tissue culture and biotechnology for the genetic improvement of watermelon. Plant Cell Tissue and Organ Culture, 77, 231-243. doi:10.1023/B:TICU.0000018428.43446.58
[50] Muruganantham, M., Ganapathi, A., Selvaraj, N., Anand, R.P., Vasudevan, A. and Vengadesan, G. (2002) Adenine sulphate and L-glutamine enhance multiple shoot induction from cotyledon explants of melon (Cucumis melo L. cv. Swarna). Cucurbit Genetics Cooperative Report, 25, 22-24.
[51] Compton, M.E. and Gray, D.J. (1992) Micropropagation as a means of rapidly propagating triploid and tetraploid watermelon. Proceedings of the Florida State Horticultural Society, 105, 352-354.
[52] Compton, M.E., Gray, D.J. and Elmstrom, G.W. (1993) A simple protocol for micropropagating diploid and tetraploid watermelon using shoot-tip explants. Plant Cell Tissue and Organ Culture, 33, 211-217. doi:10.1007/BF01983236
[53] Compton, M.E., Pierson, B.L. and Staub, J.E. (2001) Micropropagation for recovery of Cucumis hystrix. Plant Cell Tissue and Organ Culture, 64, 63-67. doi:10.1023/A:1010645206280
[54] Bhatia, P. and Ashwath, N. (2004) Comparative performance of micropropagated and seed-grown tomato plants. Biologia Plantarum, 48, 625-628. doi:10.1023/B:BIOP.0000047165.52040.d0
[55] Mandal, A.B., Maiti, A., Cowdhury, B. and Elanchezhian, R. (2001) Isoenzyme markers in varietal identification of banana. In Vitro Cellular and Developmental BiologyPlant, 37, 599-604. doi:10.1007/s11627-001-0105-z
[56] Peredo, E.L., Revilla, M.á. and Arroyo-García, R. (2006) Assessment of genetic and epigenetic variation in hop plants regenerated from sequential subcultures of organogenic calli. Journal of Plant Physiology, 163, 1071-1079. doi:10.1016/j.jplph.2005.09.010
[57] Smykal, P., Valledor, L., Rodríguez, R. and Griga, M. (2007) Assessment of genetic and epigenetic stability in long-term in vitro shoot culture of pea (Pisum sativum L.). Plant Cell Reports, 26, 1985-1998. doi:10.1007/s00299-007-0413-9
[58] Gagliardi, R.F., Hanai, L.R., Pacheco, G., Oliveira, C.A., Carneiro, L.A., Montenegro, V.J.F., Mansur, E. and Vieira, M.L.C. (2007) Assessment of genetic stability among in vitro plants of Arachis retusa using RAPD and AFLP markers for germplasm preservation. Journal of Integrative Plant Biology, 49, 307-312. doi:10.1111/j.1744-7909.2007.00402.x
[59] Sarmento, D., Martins, M. and Oliveira, M.M. (2005) Evaluation of somaclonal variation in almond using RAPD and ISSR markers. Options Méditerranéennes, 63, 391-395.
[60] Pina-Escutia, J.L., Vázquez-García, L.M. and Arzate-Fernández, A.M. (2010) In vitro regeneration and genetic fidelity of Tigridia pavonia (L.f.) DC. Electronic Journal of Biotechnology, 13, 1-7. doi:10.2225/vol13-issue1-fulltext-1
[61] Popescu, C.F., Falk, A. and Glimelius, K. (2002) Application of AFLPs to characterize somaclonal variation in anther-derived grapevines. Vitis, 41, 177-182.
[62] Polanco, C. and Ruiz, M.L. (2002) AFLP analysis of somaclonal variation in Arabidopsis thaliana regenerated plants. Plant Science, 162, 817-824. doi:10.1016/S0168-9452(02)00029-8
[63] Kumar, M.B., Barker, R.E. and Reed, B.M. (1999) Morphological and molecular analysis of genetic stability in micropropagated Fragaria x Ananassa cv. pocahontas. In Vitro Cellular and Developmental Biology-Plant, 35, 254-258. doi:10.1007/s11627-999-0088-8
[64] Soniya, E.V., Banerjee, N.S. and Das, M.R. (2001) Genetic analysis of somaclonal variation among callus-derived plants of tomato. Current Science, 80, 1213-1215.
[65] a

comments powered by Disqus

Copyright © 2020 by authors and Scientific Research Publishing Inc.

Creative Commons License

This work and the related PDF file are licensed under a Creative Commons Attribution 4.0 International License.