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Epicotyl sections as targets for plant regeneration and transient transformation of common bean using Agrobacterium tumefaciens

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Abstract

Common bean is recalcitrant to genetic transformation, due to limited regeneration capacity and low DNA transfer rates. The effect of different parameters on T-DNA transfer from Agrobacterium tumefaciens, was studied by measuring transient expression of the β-glucuronidase gene in Phaseolus vulgaris L. cv. CIAP7247F. Epicotyl containing seedling explants were inoculated with Agrobacterium EHA101 and C58C1RifR(pMP90) strains harboring the binary vector pTJK136 with GusA gene on the T-DNA. Parameters studied were temperature and light regime during co-cultivation, explant injury, and the acetosyringone concentration for vir gene induction. The co-cultivation temperature and photoperiod had a significant effect on Agrobacterium DNA transfer. In addition, explant injury and supplementation of the co-cultivation medium with acetosyringone increased the GUS activity. Optimal T-DNA transfer was obtained under the following conditions: co-cultivation at 25°C in darkness, injuring the explants with carborundum, and supplementation of the co-cultivation medium with 200 μM acetosyringone. This T-DNA delivery system was combined with a direct organogenesis protocol using epicotyl explants and fertile regenerants were recovered from tissue transformed with Agrobacterium. However, no transmission of transgenes to progeny could be observed, suggesting that the obtained plants were chimeras.

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References

  • Amugune N, Anyango B, Mukiama T (2011) Agrobacterium-mediated transformation of common bean. Afr Crop Sci J 19:137–147

    Google Scholar 

  • An G, Ebert PR, Mitra A, Ha SB (1988) Binary vectors. In: Gelvin SB, Schilperoort RA (eds) Plant molecular biology manual. Kluwer Academic Publishers, Great Britain, pp 1–19

    Google Scholar 

  • Angenon G, Thu TT (2011) Genetic transformation. In: Pratap A, Kumar J (eds) Biology and breeding of food legumes. CABI, Wallingford, pp 178–192

    Chapter  Google Scholar 

  • Aragão FJL, Barros LMG, Brasileiro ACM, Ribeiro S, Smith F, Sanford J, Faria J, Rech E (1996) Inheritance of foreign genes in transgenic bean (Phaseolus vulgaris L.) co-transformed via particle bombardment. Theor Appl Genet 93:142–150

    Article  PubMed  Google Scholar 

  • Arellano J, Fuentes SI, Castillo-Espana P, Hernández G (2009) Regeneration of different cultivars of common bean (Phaseolus vulgaris L.) via indirect organogenesis. Plant Cell Tissue Organ Cult 96:11–18

    Article  CAS  Google Scholar 

  • Bermúdez-Caraballoso I, Collado R, García LR, Veitía N, Torres D, Romero C, Angenon G (2007) Empleo de los agentes selectivos Geneticina G-418 e Higromicina B para la transformación genética en Phaseolus vulgaris variedad CIAP7247F. Biotecnol Veg 7:205–210

    Google Scholar 

  • Cheng ZM, Schnurr J, Kapaun J (1998) Timentin as an alternative antibiotic for suppression of Agrobacterium tumefaciens in genetic transformation. Plant Cell Rep 17:646–649

    Article  CAS  Google Scholar 

  • Collado R, Veitía N, Bermúdez-Caraballoso I, García L, Torres D, Romero C, Lorenzo J, Angenon G (2013) Efficient in vitro plant regeneration via indirect organogenesis for different common bean cultivars. Sci Hortic 153:109–116

    Article  CAS  Google Scholar 

  • De Clercq J, Zambre M, Van Montagu M, Dillen W, Angenon G (2002) An optimized Agrobacterium-mediated transformation procedure for Phaseolus acutifolius A. Gray. Plant Cell Rep 21:333–340

    Article  Google Scholar 

  • Delgado-Sánchez P, Saucedo-Ruiz M, Guzmán-Maldonado S, Villordo-Pineda E, González-Chavira M, Fraire-Velázquez S, Acosta-Gallegos J, Mora-Avilés A (2006) An organogenic plant regeneration system for common bean (Phaseolus vulgaris L.). Plant Sci 170:822–827

  • Dillen W, De Clercq J, Goossens A, Van Montagu M, Angenon G (1997) Agrobacterium-mediated transformation of Phaseolus acutifolius A. Gray. Theor Appl Genet 94:151–158

    Article  CAS  Google Scholar 

  • Faize M, Faize L, Burgos L (2010) Using quantitative real-time PCR to detect chimeras in transgenic tobacco and apricot and to monitor their dissociation. BMC Biotechnol 10:53–60

    Article  PubMed  PubMed Central  Google Scholar 

  • Fullner KJ, Nester EW (1996) Temperature affects the T-DNA transfer machinery of Agrobacterium tumefaciens. J Bacteriol 178:1498–1504

    CAS  PubMed  PubMed Central  Google Scholar 

  • Gatica Arias AM, Muñoz Valverde J, Ramírez Fonseca P, Valdez Melara M (2010) In vitro plant regeneration system for common bean (Phaseolus vulgaris): effect of N6-benzylaminopurine and adenine sulphate. Electron J Biotechnol 13:6–7

    Article  Google Scholar 

  • Gupta S, Bhat V, Gupta M (2006) Somatic embryogenesis and Agrobacterium mediated genetic transformation in Indian accessions of lucerne (Medicago sativa L.). Indian J Biotechnol 5:269–275

    CAS  Google Scholar 

  • Heinz DJ, Mee GWP (1969) Plant differentiation from callus tissue of Saccharum species. Crop Sci 9:346–348

    Article  Google Scholar 

  • Hoa TTC, Tran VH, La Cao T (2008) Transformation efficiencies of the soybean variety PC 19 [Glycine max (L.) Merril] using Agrobacterium tumefaciens and the cotyledonary node method. Omonrice 16:1–8

    Google Scholar 

  • Hong HP, Zhang H, Olhoft P, Hill S, Wiley H, Toren E, Hillebrand H, Jones T, Cheng M (2007) Organogenic callus as the target for plant regeneration and transformation via Agrobacterium in soybean (Glycine max (L.) Merr.). In Vitro Cell Dev Biol-Plant 43:558–568

    Article  CAS  Google Scholar 

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

    CAS  PubMed  PubMed Central  Google Scholar 

  • Kapila J, De Rycke R, Van Montagu M, Angenon G (1997) An Agrobacterium-mediated transient gene expression system for intact leaves. Plant Sci 122:101–108

    Article  CAS  Google Scholar 

  • Kwapata K, Sabzikar R, Sticklen MB, Kelly JD (2010) In vitro regeneration and morphogenesis studies in common bean. Plant Cell Tissue Organ Cult 100:97–105

  • Kwapata K, Nguyen T, Sticklen M (2012) Genetic transformation of common bean (Phaseolus vulgaris L.) with the Gus color marker, the bar herbicide resistance, and the barley (Hordeum vulgare) HVA1 drought tolerance genes. Int J Agron. doi:10.1155/2012/198960

    Google Scholar 

  • Lippincott BB, Lippincott JA (1969) Bacterial attachment to a specific wound site as an essential stage in tumor initiation by Agrobacterium tumefaciens. J Bacteriol 97:620–628

    CAS  PubMed  PubMed Central  Google Scholar 

  • Liu Z, Park BJ, Kanno A, Kameya T (2005) The novel use of a combination of sonication and vacuum infiltration in Agrobacterium-mediated transformation of kidney bean (Phaseolus vulgaris L.) with lea gene. Mol Breed 16:189–197

    Article  CAS  Google Scholar 

  • Mendel R, Müller B, Schulze J, Kolesnikov V, Zelenin A (1989) Delivery of foreign genes to intact barley cells by high-velocity microprojectiles. Theor Appl Genet 78:31–34

    Article  CAS  PubMed  Google Scholar 

  • Mohamed MF, Cao J, Earle ED (2006) Toward production of genetically modified common bean via Agrobacterium-mediated transformation. Ann Rep-Bean Improv Coop 49:147–148

    Google Scholar 

  • Mukeshimana G, Ma Y, Walworth AE, Song G, Kelly JD (2013) Factors influencing regeneration and Agrobacterium tumefaciens-mediated transformation of common bean (Phaseolus vulgaris L.). Plant Biotechnol Rep 7:59–70

    Article  Google Scholar 

  • Murashige T, Skoog F (1962) A revised medium for rapid growth and bio assays with tobacco tissue cultures. Physiol Plant 15:473–497

    Article  CAS  Google Scholar 

  • Oberpichler I, Rosen R, Rasouly A, Vugman M, Ron EZ, Lamparter T (2008) Light affects motility and infectivity of Agrobacterium tumefaciens. Environ Microbiol 10:2020–2029

    Article  CAS  PubMed  Google Scholar 

  • Opabode JT (2006) Agrobacterium-mediated transformation of plants: emerging factors that influence efficiency. Biotechnol Mol Biol Rev 1:12–20

    Google Scholar 

  • Quintero-Jiménez A, Espinosa-Huerta E, Acosta-Gallegos J, Guzmán-Maldonado H, Mora-Avilés M (2010) Enhanced shoot organogenesis and regeneration in the common bean (Phaseolus vulgaris L.). Plant Cell Tissue Organ Cult 102:381–386

    Article  Google Scholar 

  • Rech EL, Vianna GR, Aragao FJL (2008) High-efficiency transformation by biolistics of soybean, common bean and cotton transgenic plants. Nat Protoc 3:410–418

    Article  CAS  PubMed  Google Scholar 

  • Riker A (1926) Studies on the influence of some environmental factors on the development of crown gall. J Agric Res 32:83–96

    Google Scholar 

  • Riva G, González-Cabrera J, Vásquez-Padrón R, Ayra-Pardo C (1998) Agrobacterium tumefaciens: a natural tool for plant transformation. Electron J Biotechnol 1:1–6

    Google Scholar 

  • Sen P, Chatterjee G, Kumar P, Sen S (1986) Enhancement of attachment of Agrobacterium tumefaciens to plant cell surface results in increase in genetic transformation. Indian J Exp Biol 24:153–155

    CAS  Google Scholar 

  • Stachel SE, Messens E, Van Montagu M, Zambryski P (1985) Identification of the signal molecules produced by wounded plant cells that activate T-DNA transfer in Agrobacterium tumefaciens. Nature 318:624–629

    Article  Google Scholar 

  • Thu TT, Xuan Mai TT, Dewaele E, Farsi S, Tadesse Y, Angenon G, Jacobs M (2003) In vitro regeneration and transformation of pigeonpea [Cajanus cajan (L.) Millsp]. Mol Breed 11:159–168

    Article  CAS  Google Scholar 

  • Varisai Mohamed S, Sung JM, Jeng TL, Wang CS (2006) Organogenesis of Phaseolus angularis L.: high efficiency of adventitious shoot regeneration from etiolated seedlings in the presence of N6-benzylaminopurine and thidiazuron. Plant Cell Tissue Organ Cult 86:187–199

    Article  CAS  Google Scholar 

  • Veltcheva M, Svetleva D, Petkova S, Perl A (2005) In vitro regeneration and genetic transformation of common bean (Phaseolus vulgaris L.)—problems and progress. Sci Hortic 107:2–10

    Article  CAS  Google Scholar 

  • Villemont E, Dubois F, Sangwan RS, Vasseur G, Bourgeois Y, Sangwan-Norreel BS (1997) Role of the host cell cycle in the Agrobacterium-mediated genetic transformation of Petunia: evidence of an S-phase control mechanism for T-DNA transfer. Planta 201:160–172

    Article  CAS  Google Scholar 

  • Zambre M, Goossens A, Cardona C, Van Montagu M, Terryn N, Angenon G (2005) A reproducible genetic transformation system for cultivated Phaseolus acutifolius (tepary bean) and its use to assess the role of arcelins in resistance to the Mexican bean weevil. Theor Appl Genet 110:914–924

    Article  CAS  PubMed  Google Scholar 

  • Zambre M, Terryn N, De Clercq J, De Buck S, Dillen W, Van Montagu M, Van Der Straeten D, Angenon G (2003) Light strongly promotes gene transfer from Agrobacterium tumefaciens to plant cells. Planta 216:580–586

    CAS  PubMed  Google Scholar 

  • Zhang Z, Coyne DP, Mitra A (1997) Factors affecting Agrobacterium-mediated transformation of common bean. J Am Soc Hortic Sci 122:300–305

    CAS  Google Scholar 

  • Zupan J, Muth TR, Draper O, Zambryski P (2000) The transfer of DNA from Agrobacterium tumefaciens into plants: a feast of fundamental insights. Plant J 23:11–28

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

This research was financially supported by the Institutional University Collaboration programme with Universidad Central “Marta Abreu” de Las Villas from the Flemish Interuniversity Council (VLIR IUC UCLV). Thanks also go to Edilio Quintero from CIAP for kindly supplying seeds of common bean cv. CIAP7247F.

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Authors and Affiliations

  1. Instituto de Biotecnología de las Plantas, Universidad Central “Marta Abreu” de Las Villas, Carretera a Camajuaní km 5.5, Santa Clara, Villa Clara, PC-54 830, Cuba

    R. Collado, I. Bermúdez-Caraballoso, L. R. García, N. Veitía, D. Torres & C. Romero

  2. Laboratory of Plant Genetics, Vrije Universiteit Brussel (VUB), Pleinlaan 2, B-1050, Brussels, Belgium

    R. Collado & G. Angenon

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  1. R. Collado
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  2. I. Bermúdez-Caraballoso
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  3. L. R. García
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  4. N. Veitía
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  5. D. Torres
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  6. C. Romero
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  7. G. Angenon
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Correspondence to R. Collado.

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Editor: Ewen Mullins

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Collado, R., Bermúdez-Caraballoso, I., García, L.R. et al. Epicotyl sections as targets for plant regeneration and transient transformation of common bean using Agrobacterium tumefaciens . In Vitro Cell.Dev.Biol.-Plant 52, 500–511 (2016). https://doi.org/10.1007/s11627-016-9769-2

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