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An efficient Agrobacterium tumefaciens-mediated genetic transformation of “Egusi” melon (Colocynthis citrullus L.)

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Abstract

Cotyledonary explants of two “Egusi” genotypes, ‘Ejagham’ and NHC1-130, were co-cultivated with Agrobacterium tumefaciens strain EHA101 carrying either plasmid pIG121-Hm harbouring genes coding for beta-glucuronidase (gus), hygromycin phosphotransferase (hpt) and neomycin phosphotransferase II (nptII) or plasmid pBBRacdS harbouring these same genes along with a gene coding for 1-aminocyclopropane-1-carboxylate (ACC) deaminase. Six weeks after co-cultivation, more than 35% of explants produced shoots in both cultivars. A DNA fragment corresponding to the gus gene or the selection marker nptII was amplified from genomic DNA extracted from leaves of regenerated plant clones rooted on hormone-free MS medium containing 100 mg/l kanamycin, suggesting their transgenic nature. Southern blot analysis confirmed successful integration of one to three copies of the gus gene. Transformation efficiencies of cultivar NHC1-130 with EHA101(pIG121-Hm) and EHA101(pIG121-Hm, pBBRacdS) were 3.8% and 10%, respectively, which were higher than those obtained for cultivar ‘Ejagham’ of 2.4% and 5.7%, respectively. Co-cultivation medium containing 5 mg/l BA was effective for obtaining high transformation efficiency for both cultivars as compared with that without it.

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Abbreviations

BA:

Benzylaminopurine

GUS:

β-Glucuronidase

MS:

Murashige and Skoog

NPTII:

Neomycin phosphotransferase

PCR:

Polymerase chain reaction

References

  • Castelblanque L, Marfa V, Claveria E, Martinez I, Perez-Grau L, Dolcet-Sanjuan R (2008) Improving the genetic transformation efficiency of Cucumis melo subsp. melo ‘Piel de Sapo’ via Agrobacterium. Cucurbitaceae 2008. In: Pitrat M (ed) Proceedings of the IXth EUCARPIA meeting on genetic and breeding of Cucurbitaceae. INRA, Avignon (France), May 21–24th, 2008

  • Chovelon V, Restier V, Dogimont C, Aarrout J (2008) Histological study of shoot organogenesis in melon (Cucumis melo L.) after genetic transformation. Cucurbitaceae 2008. In: Pitrat M (ed) Proceedings of the IXth EUCARPIA meeting on genetic and breeding of Cucurbitaceae. INRA, Avignon (France), May 21–24th, 2008

  • Cuozzo M, O’Connell KM, Kaniewski W, Fang R-X, Chua N-H, Tumer N (1988) Viral protection in transgenic tobacco plants expressing the cucumber mosaic virus coat protein or its antisense RNA. Nat Biotechnol 6:549–557

    Article  CAS  Google Scholar 

  • Curuk S, Centiner S, Elman C, Xia X, Wang Y, Yeheskel A, Zilberstein L, Perl-Treves R, Watad A, Gaba V (2005) Transformation of recalcitrant melon (Cucumis melo L.) cultivars is facilitated by wounding with carborundum. Eng Life Sci 5:169–177

    Article  Google Scholar 

  • Dabuaza M, Bordeas M, Salvador A, Roig L-A, Moreno V (1997) Plant regeneration and Agrobacterium-mediated transformation of cotyledon explants of Citrullus colocynthis (L.) Schard. Plant Cell Rep 16:888–892

    Article  Google Scholar 

  • Davis WE, Miller AR, Linerberger RD (1992) Studies on the effects of ethylene on transformation of tomato cotyledons (Lycopersicon esculentum Mill.) by Agrobacterium tumefaciens. J Plant Physiol 139:302–309

    Google Scholar 

  • Ezura H, Yuhashi KI, Yasuta T, Minamisawa K (2000) Effect of ethylene on Agrobacterium tumefaciens-mediated gene transfer to melon. Plant Breed 119:75–79

    Article  CAS  Google Scholar 

  • Fang G, Grumet R (1990) Agrobacterium tumefaciens mediated transformation and regeneration of musk melon plants. Plant Cell Rep 9:160–164

    Article  CAS  Google Scholar 

  • Gaba V, Less H, Antigus Y (1992) Transformation of melon by particle acceleration. Plant Physiol 99(Suppl):137

    Google Scholar 

  • Galperin M, Patlis L, Ovadia A, Wolf D, Zelcer A, Kenigsbuch D (2003) A melon genotype with superior competence for regeneration and transformation. Plant Breed 122:66–69

    Article  Google Scholar 

  • Gasser CS, Fraley RT (1989) Genetically engineering plants for crop improvement. Science 244:1293–1331

    Article  CAS  PubMed  Google Scholar 

  • Giwa SO, Chuah LA, Adam NM (2009) Optimization of the production of biodiesel from ‘Egusi’ melon (Colocynthis citrullus L.) oil using response surface methodology. International Advance of Technology Congress (ATC), PWTC, Malaysia, Article No. 028, November 3–5, 2009

  • Guis M, Ben-Amor M, Latche A, Peche JC, Roustan JP (2000) A reliable system for the transformation of cantaloupe charentais melon (Cucumis melo L. var. cantaloupensis) leading to a majority of diploid regenerants. Sci Hort 84:91–99

    Article  CAS  Google Scholar 

  • Gusmini G, Wehner TC, Jarret RL (2004) Inheritance of Egusi seed type in watermelon. J Hered 95:268–270

    Article  CAS  PubMed  Google Scholar 

  • Hamill JD, Rounsley S, Spencer A, Todd G, Rhodes MJC (1991) The use of the polymerase chain reaction in plant transformation studies. Plant Cell Rep 10:221–224

    Article  CAS  Google Scholar 

  • Han JS, Kim KC, Park SH, Hirschi DK, Mok IG (2005) Agrobacterium-mediated transformation of bottle gourd (Lagenaria siceraria Standl.). Plant Cell Rep 23:692–698

    Article  CAS  PubMed  Google Scholar 

  • Hood EE, Helmer GL, Fraley RT, Chilton MD (1986) The hypervirulence of Agrobacterium tumefaciens A281 is encoded in a region of pTiBo54 outside TDNA. J Bacteriol 168:1291–1301

    CAS  PubMed  Google Scholar 

  • Jefferson RK, Kavanagh TA, Bevan MW (1987) GUS fusion: beta-glucuronidase as a sensitive and versatile gene fusion marker in higher plants. EMBO J 6:3901–3907

    CAS  PubMed  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bioassay with tobacco tissue cultures. Physiol Plant 15:437–497

    Article  Google Scholar 

  • Nonaka S, Sugawara M, Minamisawa K, Yuhashi K, Ezura H (2008a) 1-Aminocyclopropane-1-carboxylate deaminase enhances Agrobacterium tumefaciens-mediated gene transfer into plant cell. Appl Env Microbiol 74:2526–2528

    Article  CAS  Google Scholar 

  • Nonaka S, Yuhashi K, Takada K, Sugawara M, Minamisawa K, Ezura H (2008b) Ethylene production in plants during gene transformation suppresses vir gene expression in Agrobacterium tumefaciens. New Phytol 178(3):647–656

    Article  CAS  PubMed  Google Scholar 

  • Nora FR, Peters JA, Schuch MW, Lucchetta L, Marini L, Jorge A, Rombaldi CV (2001) Melon regeneration and transformation using an apple ACC oxidase antisense gene. Rev Bras de Agrociencia 7:201–204

    Google Scholar 

  • Ntui VO, Uyoh EA (2005) Inheritance of stripe pattern on fruits and seed colour in “Egusi” melon (Colocynthis citrullus L.). Glob J Agric Sci 4:29–32

    Google Scholar 

  • Ntui VO, Thirukkumaran G, Iioka S, Mii M (2009) Efficient plant regeneration via organogenesis in “Egusi” melon (Colocynthis citrullus L.). Sci Hort 119:397–402

    Article  CAS  Google Scholar 

  • Ogawa Y, Mii M (2007) Meropenem and moxalactam: novel β-lactam antibiotics for efficient Agrobacterium-mediated transformation. Plant Sci 172:564–572

    Article  CAS  Google Scholar 

  • Ohta S, Mira S, Hattori T, Nakamura K (1990) Construction and expression in tobacco of a β-glucuronidase (GUS) reporter gene containing an intron within the coding sequence. Plant Cell Physiol 31:805–813

    CAS  Google Scholar 

  • Oyulu C (1977) A quantitative and qualitative study of seed type in ‘Egusi’ (Colocynthis citrullus L.). Trop Sci 19:55–62

    Google Scholar 

  • Powell-Abel P, Nelson RS, De B, Hoffmann N, Rogers SG, Fraley RT, Beachy RN (1986) Delay of disease development in transgenic plants that express the tobacco mosaic virus coat protein gene. Science 232:738–743

    Article  Google Scholar 

  • Purseglove JW (1991) Tropical crops (dicotyledons). Longman, New York

    Google Scholar 

  • Rhimi A, Hernould M, Boussaid M (2007) Agrobacterium-mediated transformation of Tunisian Cucumis melo cv. Maazoun. Afr J Biotechnol 6(18):2162–2165

    CAS  Google Scholar 

  • Rogers OS, Bendich JA (1988) Extraction of DNA from plant tissues. In: Gelvin SB, Schiliperoort RA, Verma DPS (eds) Plant molecular biology manual, vol A6. Kluwer, Dordrecht, pp 1–10

    Google Scholar 

  • Schippers RR (2000) African indigenous vegetables. An overview of the cultivated species. Natural Resources Institute/ACP-EU Technical Centre for Agricultural and Rural Cooperation, Chatham

    Google Scholar 

  • Schippers RR (2004) Cucurbitaceae. In: Légumes africains indigènes: présentation des espèces cultivées. Margraf Publishers/CTA, Wuerzburg, pp 113–182

  • Valles MP, Lasa JM (1994) Agrobacterium- mediated transformation of commercial melon (Cucumis melo L., cv Amarilloo Oro). Plant Cell Rep 13:145–148

    Article  CAS  Google Scholar 

  • Vodouhe RS, Achigan-Dako GE, Adjakidje V (2001) Observation de la diversité génétique des Egusi collectées au Bénin et au Togo. In: Agbo BP et al. (eds) Actes 2 de l’atelier scientifique sud et centre, 12–13 décembre 2001, Institut National des Recherches Agricoles du Bénin. Niaouli, INRAB, Bénin, pp 53–61

  • Wu HW, Yu TA, Raja JAJ, Wang HC, Yeh SD (2009) Generation of transgenic oriental melon resistant to Zucchini yellow mosaic virus by an improved cotyledon-cutting method. Plant Cell Rep 28:1053–1064

    Article  CAS  PubMed  Google Scholar 

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Authors and Affiliations

  1. Laboratory of Plant Cell Technology, Graduate School of Horticulture, Chiba University, 648 Matsudo, Matsudo, Chiba, 271-8510, Japan

    Valentine Otang Ntui, Raham Sher khan, Dong Poh Chin, Ikuo Nakamura & Masahiro Mii

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  1. Valentine Otang Ntui
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  2. Raham Sher khan
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  3. Dong Poh Chin
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  4. Ikuo Nakamura
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  5. Masahiro Mii
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Correspondence to Masahiro Mii.

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Ntui, V.O., khan, R.S., Chin, D.P. et al. An efficient Agrobacterium tumefaciens-mediated genetic transformation of “Egusi” melon (Colocynthis citrullus L.). Plant Cell Tiss Organ Cult 103, 15–22 (2010). https://doi.org/10.1007/s11240-010-9748-y

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