Skip to main content
SpringerLink
Log in

Factors influencing regeneration and Agrobacterium tumefaciens-mediated transformation of common bean (Phaseolus vulgaris L.)

  • Original Article
  • Published:
Plant Biotechnology Reports Aims and scope Submit manuscript

Abstract

A systematic study was carried out to optimize regeneration and Agrobacterium tumefaciens-mediated transformation of four common bean (Phaseolus vulgaris L.) cultivars; Red Hawk, Matterhorn, Merlot, and Zorro, representing red kidney, great northern, small red, and black bean commercial classes, respectively. Regeneration capacity of leaf explants, stem sections, and embryo axes were evaluated on 30 media each containing Murashige and Skoog (MS) medium and different combinations of plant growth regulators. For stem sections and leaf explants, none of the media enabled plant regeneration from any of the four cultivars tested, indicating the recalcitrance of bean regeneration from these tissues. In contrast, several media enabled multiple shoot production from embryo axis explants, although optimal regeneration media was genotype-dependent. Under optimal regeneration conditions, multiple shoots, 2.3–10.8 on average for each embryogenic explant, were induced from embryo axis explants at frequencies of 93 % for ‘Merlot’, 80 % for ‘Matterhorn’, 73 % for ‘Red Hawk’, and 67 % for ‘Zorro’. Transient expression studies monitored by an intron-interrupted gusA on explants transformed with A. tumefaciens strains GV3101, LBA4404, and EHA105 indicated that all three A. tumefaciens strains tested were efficient in gene delivery. Gene delivery depended on parameters including strain of A. tumefaciens, co-cultivation time, explant type, and bean genotype. Agroinfiltration also enhanced gene delivery. Kanamycin-resistant and GUS-positive calluses were induced from leaf, stem, and embryo axis explants. Chimeric transformants were obtained from embryo axis explants and showed partial GUS-staining. Lack of efficient regeneration from non-meristem containing tissues is the main limitation for stable transformation of common bean.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  • Aragão FJL, de Sá MFG, Almeida ER, Gander ES, Rech EL (1992) Particle bombardment-mediated transient expression of a Brazil nut methionine-rich albumin in bean (Phaseolus vulgaris L.). Plant Mol Biol 20:357–359

    Article  PubMed  Google Scholar 

  • Aragão FJL, Ribeiro SG, Barros LMG, Brasileiro ACM, Maxwell DP, Rech EL, Faria JC (1998) Transgenic beans (Phaseolus vulgaris L.) engineered to express viral antisense RNAs showed delayed and attenuated symptoms to bean golden mosaic geminivirus. Mol Breed 4:491–499

    Article  Google Scholar 

  • Arellano J, Fuentes SI, Castillo-España P, Hernández G (2009) Regeneration of different cultivars of common bean (Phaseolus vulgaris L.) via indirect organogenesis. Plant Cell Tissue Organ Cult 96:1–11

    Article  Google Scholar 

  • Bakshi S, Sadhukhan A, Mishra S, Sahoo L (2011) Improved Agrobacterium-mediated transformation of cowpea via sonication and vacuum infiltration. Plant Cell Rep 30:2281–2292

    Article  PubMed  CAS  Google Scholar 

  • Bonfim K, Faria JC, Nogueira EOPL, Mendes ÉA, Aragão FJL (2007) RNAi-mediated resistance to Bean golden mosaic virus in genetically engineered common bean (Phaseolus vulgaris). Mol Plant Microbe Interact 20:717–726

    Article  PubMed  CAS  Google Scholar 

  • Brasiliero ACM, Aragao FJL, Rossi S, Dusi DMA, Barros LMG, Rech EL (1996) Susceptibility of common beans to Agrobacterium ssp. Strains and improvement of Agrobacterium transformation using microprojectile bombardment. J Am Soc Hortic Sci 121:810–815

    Google Scholar 

  • Broughton WJ, Hernandez G, Blair M, Beebe S, Gepts P, Vanderleyden J (2003) Beans (Phaseolus spp.)—model food legumes. Plant Soil 252:55–128

    Article  CAS  Google Scholar 

  • Cheng M, Fry JE, Pang S, Zhou H, Hironoka CM, Duncan DR, Conner TW, Wan Y (1997) Genetic transformation of wheat mediated by Agrobacterium tumefaciens. Plant Physiol 115:971–980

    PubMed  CAS  Google Scholar 

  • Christou P (1997) Biotechnology applied to grain legumes. Field Crops Res 53:83–97

    Article  Google Scholar 

  • Crocomo OJ, Peters JE, Sharp WR (1976) Interactions of phytohormones on the control of growth and root morphogenesis in cultured Phaseolus vulgaris leaf explants. Turrialba 26:232–236

    CAS  Google Scholar 

  • Cruz de Carvalho MN, Van Le B, Zuili-Fodil Y, Pham Thi AT, Van TranThan K (2000) Efficient whole plant regeneration of common bean (Phaseolus vulgaris L.) using thin cell layer culture and silver nitrate. Plant Sci 159:223–232

    Article  PubMed  CAS  Google Scholar 

  • Dang W, Wei ZM (2009) High frequency plant regeneration from the cotyledonary node of common bean. Biol Plant 53(2):312–316

    Article  CAS  Google Scholar 

  • Davis ME, Miller AR, Lineberger RD (1991) Temporal competence for transformation of Lycopersicon esculentum (L. Mill) cotyledons by Agrobacterium tumefaciens. Relation to wound healing and soluble plant factors. J Exp Bot 42:359–364

    Article  Google Scholar 

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

    Article  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Dita MA, Rispail N, Prats E, Rubiales D, Singh KB (2006) Biotechnology approaches to overcome biotic and abiotic stress constraints in legumes. Euphytica 147:1–24

    Article  Google Scholar 

  • Estrada-Navarrete G, Alvarado-Affantranger X, Olivares JE, Díaz-Camino Cl, Santana O, Murillo E, Guillén G, Sánchez-Guevara N, Acosta J, Quinto C, Li D, Gresshoff PM, Sánchez F (2006) Agrobacterium rhizogenes transformation of the Phaseolus spp.: a tool for functional genomics. Mol Plant Microbe Interact 19:1385–1393

    Article  PubMed  CAS  Google Scholar 

  • Gamborg OL, Miller RA, Ojima K (1968) Nutrient requirements of suspension cultures of soybean root cells. Exp Cell Res 50:151–158

    Article  PubMed  CAS  Google Scholar 

  • Gatica Arias AM, Valverde JM, Fonseca PR, Melara MV (2010) In vitro plant regeneration system for common bean (Phaseolus vulgaris): effect of N6-benzylaminopurine and adenine sulphate. Electronic J Biotechnol 13:1–8

    Google Scholar 

  • Geerts P, Mergeai G, Baudon JP (2000) Succeeded plant regeneration from very immature embryos of Phaseolus vulgaris. Ann Rep Bean Improv Coop 43:206–207

    Google Scholar 

  • Grant J, Cooper P (2006) Peas (Pisum sativum L.). Methods Mol Biol 343:337–345

    Google Scholar 

  • Hiei Y, Ohta S, Komari T, Kumashiro T (1994) Efficient transformation of rice (Oryza sativa L.) mediated by Agrobacterium and sequence analysis of the boundaries of the T-DNA. Plant J 6:271–282

    Article  PubMed  CAS  Google Scholar 

  • Hoekema A, Hirsch PR, Hooykaas PJJ, Schilperoort RA (1983) A binary plant vector strategy based on separation of vir- and T-region of the Agrobacterium tumefaciens Ti-plasmid. Nature 303:179–180

    Article  CAS  Google Scholar 

  • Hood EA, Gelvin SB, Melchers LS, Hoekema A (1993) New Agrobacterium helper plasmids for gene transfer to plants. Transgenic Res 2:208–218

    Article  CAS  Google Scholar 

  • Huda S, Islam R, Bari MA, Asaduzzaman M (2003) Shoot differentiation from cotyledon derived callus of chickpea (Cicer arietinum L.). Plant Cell Tissue Organ Cult 13:53–59

    Google Scholar 

  • Indurker S, Misra HS, Eapen S (2010) Agrobacterium-mediated transformation in chickpea (Cicer arietinum L.) with an insecticidal protein gene: optimization of different factors. Physiol Mol Biol Plants 16:273–284

    Article  CAS  Google Scholar 

  • Jayanand B, Sudarsanam G, Sharma KK (2003) An efficient protocol for the regeneration of whole plants of chickpea (Cicer arietinum L.) by using axilary meristem explants derived from in vitro-germinated seeds. In Vitro Cell Dev Biol Plant 39:171–179

    Article  Google Scholar 

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

    PubMed  CAS  Google Scholar 

  • Kim JW, Minamikawa T (1996) Transformation and regeneration of French bean plants by the particle bombardment process. Plant Sci 117:131–138

    Article  CAS  Google Scholar 

  • Koncz C, Schell J (1986) The promoter of the TL-DNA gene 5 controls the tissue-specific expression of chimeric genes carried by a novel type of Agrobacterium binary vector. Mol Gen Genet 204:383–396

    Article  CAS  Google Scholar 

  • Krishna G, Reddy PS, Ramteke PW, Bhattacharya PS (2010) Progress of tissue culture and genetic transformation research in pigeon pea [Cajanus cajan (L.) Millsp.]. Plant Cell Rep 29:1079–1095

    Article  PubMed  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

    Article  CAS  Google Scholar 

  • Lewis ME, Bliss FA (1994) Tumor formation and glucuronidase expression in Phaseolus vulgaris inoculated with Agrobacterium tumefaciens. J Am Soc Hortic Sci 119:361–366

    CAS  Google Scholar 

  • Li HQ, Sautter C, Potrykus I, Puonti-Kaerlas J (1996) Genetic transformation of cassava (Manihot esculenta Crantz). Nat Biotechnol 14:736–740

    Article  PubMed  CAS  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 

  • Lloyd G, Mccown B (1980) Commercially feasible micropropagation of mountain laurel, Kalmia latifolia, by use of shoot tip culture. Proc Int Plant Prop Soc 30:421–427

    Google Scholar 

  • Mahmoudian M, Yucel M, Oktem HA (2002) Transformation of lentil (Lens culinaris M.) cotyledonary nodes by vacuum infiltration of Agrobacterium tumefaciens. Plant Mol Biol Rep 20:251–257

    Article  Google Scholar 

  • Malik KA, Saxena PK (1991) Regeneration in Phaseolus vulgaris L. Promotive role of N6-benzylaminopurine in cultures from juvenile leaves. Planta 184:148–150

    Article  CAS  Google Scholar 

  • Mariotti D, Fontana GS, Santini L (1989) Genetic transformation of grain legumes: Phaseolus vulgaris L. and P. coccineus L. J Genet Breed 43:77–82

    Google Scholar 

  • McClean P, Chee P, Held B, Simental J, Drong RF, Slightom J (1991) Susceptibility of dry bean (Phaseolus vulgaris L.) to Agrobacterium infection: transformation of cotyledonary and hypocotyl tissues. Plant Cell Tissue Organ Cult 24:131–138

    Article  Google Scholar 

  • Mehrotra M, Sanyal I, Amla DV (2011) High-efficiency Agrobacterium-mediated transformation of chickpea (Cicer arietinum L.) and regeneration of insect-resistant transgenic plants. Plant Cell Rep 30:1603–1616

    Article  PubMed  CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Muthukumar B, Mariamma M, Veluthambi K, Gnanam A (1996) Genetic transformation of cotyledon explants of cowpea (Vigna unguiculata L. Walp) using Agrobacterium tumefaciens. Plant Cell Rep 15:980–985

    CAS  Google Scholar 

  • Ni M, Cui D, Einstein J, Narasimhulu S, Vergara CE, Gelvin SB (1995) Strength and tissue specificity of chimeric promoters derived from the octopine and mannopine synthase genes. Plant J 7:661–676

    Article  CAS  Google Scholar 

  • Paz MM, Martinez JC, Kalvig AB, Fonger TM, Wang K (2006) Improved cotyledonary node method using an alternative explant derived from mature seed for efficient Agrobacterium-mediated soybean transformation. Plant Cell Rep 25:206–213

    Article  PubMed  CAS  Google Scholar 

  • Polowick PL, Balisk DS, Mahon JD (2004) Agrobacterium tumefaciens-mediated transformation of chickpea (Cicer arietinum L.): gene integration, expression and inheritance. Plant Cell Rep 23:485–491

    Article  PubMed  CAS  Google Scholar 

  • Popelka JC, Gollasch S, Moore A, Molving L, Higgins TJV (2006) Genetic transformation of cowpea (Vigna unguiculata L.) and stable transmission of the transgenes to progeny. Plant Cell Rep 25:304–312

    Article  PubMed  CAS  Google Scholar 

  • Quintero-Jimenez A, Espinosa-Huerta E, Acosta-Gallegos JA, Guzman-Maldonado HS, Mora-Aviles MA (2010) Enhanced shoot organogenesis and regeneration in the common bean (Phaseolus vulgaris L.). Plant Cell Tissue Organ Cult 102:381–386

    Article  Google Scholar 

  • Quoirin M, Lepoivre P (1977) Improved media for in vitro culture of Prunus sp. Acta Hort 78:437–442

    Google Scholar 

  • Russel DR, Wallace KM, Bathe JH, Martinell BJ, McCabe DE (1993) Stable transformation of PhaseoIus vulgaris via electric-discharge mediated particle acceleration. Plant Cell Rep 12:165–169

    Article  Google Scholar 

  • Saunders JW, Hosfield GL, Levi A (1987) Morphogenetic effect of 2,4-dichlorophenoxyacetic acid on pinto bean (Phaseolus vulgaris L.) leaf explants in vitro. Plant Cell Rep 6:46–49

    Article  CAS  Google Scholar 

  • Senthil G, Williamson B, Dinkins RD, Ramsay G (2004) An efficient transformation system for chickpea (Cicer arietinum L.). Plant Cell Rep 23:297–303

    Article  PubMed  CAS  Google Scholar 

  • Sharma KK, Pooja BM (2006) Peanut (Arachis hypogaea L.). In: Wang K (ed) Methods in molecular biology, vol 343: Agrobacterium Protocols, 2nd edn. vol 1. Humana Press, Totowa, pp 347–358

  • Solleti SK, Bakshi S, Purkayastha J, Panda SK, Sahoo L (2008) Transgenic cowpea (Vigna unguiculata) seeds expressing a bean a-amylase inhibitor confer resistance to storage pests, bruchid beetles. Plant Cell Rep 27:1841–1850

    Article  PubMed  CAS  Google Scholar 

  • Somers DA, Samac DA, Olhoft PM (2003) Recent advances in legume transformation. Plant Physiol 131:892–899

    Article  PubMed  CAS  Google Scholar 

  • Svetleva D, Velcheva M, Bhowmik G (2003) Biotechnology as a useful tool in common bean (Phaseolus vulgaris L.) improvement. Euphytica 131:189–200

    Article  CAS  Google Scholar 

  • Uranbey S, Sevimay CS, Kaya MD, Ipek A, Sancak C, Başalma D, Er C, Ozcan S (2005) Influence of different co-cultivation temperatures, periods and media on Agrobacterium tumefaciens-mediated gene transfer. Biol Plant 49:53–57

    Article  Google Scholar 

  • Veltcheva M, Svetleva D, Petkova SP, 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 

  • Vervliet G, Holsters M, Tecuchy H, van Montagu M, Schell J (1975) Characterization of different plaque-forming and defective temperate phages in Agrobacterium strains. J Gen Virol 26:33–48

    Article  PubMed  CAS  Google Scholar 

  • Zambre MA, De Clercq J, Vranová E, Van Montagu M, Angenon G, Dillen W (1998) Plant regeneration from embryo-derived callus in Phaseolus vulgaris L. (common bean) and P. acutifolius A. Gray (tepary bean). Plant Cell Rep 17:626–630

    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  PubMed  CAS  Google Scholar 

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

    CAS  Google Scholar 

Download references

Acknowledgments

This research was supported in part by the USDA-AFRI, PULSE CRSP and MSU Project GREEEN (Generating Research and Extension to Meet Economic and Environmental Needs).

Author information

Authors and Affiliations

  1. Department of Crop and Soil Sciences, Michigan State University, 1066 Bogue St., East Lansing, MI, 48824, USA

    Gerardine Mukeshimana & James D. Kelly

  2. Department of Horticulture, Plant Biotechnology Resource and Outreach Center, Michigan State University, 1066 Bogue St., East Lansing, MI, 48824, USA

    Yumin Ma, Aaron E. Walworth & Guo-qing Song

Authors
  1. Gerardine Mukeshimana
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yumin Ma
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Aaron E. Walworth
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Guo-qing Song
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. James D. Kelly
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to James D. Kelly.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Mukeshimana, G., Ma, Y., Walworth, A.E. et al. Factors influencing regeneration and Agrobacterium tumefaciens-mediated transformation of common bean (Phaseolus vulgaris L.). Plant Biotechnol Rep 7, 59–70 (2013). https://doi.org/10.1007/s11816-012-0237-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s11816-012-0237-0

Keywords

Search

Navigation