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Agrobacterium-mediated transformation of apple (Malus x domestica Borkh.): an assessment of factors affecting regeneration of transgenic plants

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Abstract

We have previously developed a protocol for efficient gene transfer and regeneration of transgenic calli following cocultivation of apple (cv. Jonagold) explants with Agrobacterium tumefaciens (De Bondt et al. 1994, Plant Cell Reports 13: 587–593). Now we report on the optimization of postcultivation conditions for efficient and reproducible regeneration of transgenic shoots from the apple cultivar Jonagold. Factors which were found to be essential for efficient shoot regeneration were the use of gelrite as a gelling agent and the use of the cytokinin-mimicing thidiazuron in the selective postcultivation medium. Improved transformation efficiencies were obtained by combining the hormones thidiazuron and zeatin and by using leaf explants from in vitro grown shoots not older than 4 weeks after multiplication. Attempts to use phosphinothricin acetyl transferase as a selectable marker were not successful. Using selection on kanamycin under optimal postcultivation conditions, about 2% of the leaf explants developed transgenic shoots or shoot clusters. The presence and expression of the transferred genes was verified by β-glucuronidase assays and Southern analysis. The transformation procedure has also been succesfully applied to several other apple cultivars.

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Abbreviations

BAP:

benzylaminopurine

CTAB:

hexadecyltrimethylammoniumbromide

Na2EDTA:

ethylenediamine-tetra-acetate ferric-sodium salt

FeNaEDTA:

ethylenediamine-tetra-acetate ferric-sodium salt

GA3 :

gibberellic acid 3

GusA:

β-glucuronidase

gusA:

β-glucuronidase gene of Escherichia coli

IAA:

indole acetic acid

IBA:

indole butyric acid

2iP:

N6-2-isopentenyl adenine

NAA:

naphthalene acetic acid

nptII:

neomycinphosphotransferase II gene

bar :

phosphinothricin acetyl transferase gene

PCR:

polymerase chain reaction

PPT:

phosphinothricin

STS:

silver thiosulphate

T-DNA:

transferred DNA

TDZ:

thidiazuron

X-Gluc:

5-bromo-4-chloro-3-indolyl β-D-glucuronide

Zea:

trans-Zeatin

References

  • Aldrich J, Cullis CA (1993) Plant Mol Biol Rep 11: 128–141

    Google Scholar 

  • Becker D, Kemper E, Schell J, Masterson R (1992) Plant Mol Biol 20: 1195–1197

    CAS  PubMed  Google Scholar 

  • Cornelissen M, Vandewiele M (1989) Nucleic Acids Res 17: 19–29

    Google Scholar 

  • De Bondt A, Eggermont K, Druart P, De Vil M, Goderis I, Vanderleyden J, Broekaert W (1994) Plant Cell Reports 13: 587–593

    Google Scholar 

  • Dekeyser R, Claes B, Marichal M, Van Montagu M, Caplan A (1989) Plant Physiol 90: 217–223

    Google Scholar 

  • Druart P (1980) Scientia Hortic 12: 339–342

    Google Scholar 

  • Druart P (1988) Acta Hort 227: 369–380

    Google Scholar 

  • Druart P (1990) Acta Hort 280: 117–124

    Google Scholar 

  • Escalettes V, Dosba F (1993) Plant Sci 90: 201–209

    Google Scholar 

  • Fasolo F, Zimmerman RH, Fordham I (1989) Plant Cell Tiss Org Cult 16: 75–87

    Google Scholar 

  • Hamill JD, Rounsley S, Spencer A, Todd G, Rhodes MJC (1991) Plant Cell Reports 10: 221–224

    Google Scholar 

  • Hassan MA, Swartz HJ, Inamine G, Mullineaux P (1993) Plant Cell Tiss Org Cult 33: 9–17

    Google Scholar 

  • Hood EE, Helmer GL, Frayley RT, Chilton M-D (1986) J Bacteriol 168: 1297–1301

    Google Scholar 

  • Hood EE, Gelvin SB, Melchers LS, Hoekema A (1993) Transgenic Research 2: 208–218

    Google Scholar 

  • Huetteman CA, Preece JE (1993) Plant Cell Tiss Org Cult 33: 105–119

    Google Scholar 

  • James DJ, Passey AJ, Barbara D (1990) In: Genetic engineering of crop plants, 239–248, (eds.) Lycett GW, Grierson D, Butterworths-London

    Google Scholar 

  • James DJ, Passey AJ, Rugini E (1988) J Plant Physiol 132: 148–154

    CAS  Google Scholar 

  • James DJ, Passey AJ, Barbara DJ, Bevan MW (1989) Plant Cell Reports 7: 658–661

    Google Scholar 

  • James DJ, Uratsu S, Cheng J, Negri P, Viss P, Dandekar AM (1993) Plant Cell Reports 12: 559–563

    Google Scholar 

  • Maheswaran G, Welander M, Hutchinson JF, Graham MW, Richards D (1992) J Plant Physiol 139: 560–568

    Google Scholar 

  • McBride KE, Summerfelt KR (1990) Plant Mol Biol 14: 269–276

    Google Scholar 

  • McCabe D, Christou P (1993) Plant Cell Tiss Org Cult 33: 227–236

    Google Scholar 

  • Murashige T, Skoog F (1962) Physiol Plant 15: 473–497

    CAS  Google Scholar 

  • Nichol JW, Slade D, Viss P, Stuart DA (1991) Plant Sci 79: 181–192

    Google Scholar 

  • Norelli JL, Aldwinckle HS, Destéfano-Beltran L, Jaynes JM (1994) Euphytica 77: 123–128

    Google Scholar 

  • Patat-Ochatt E, Ochatt SJ, Power JB (1988) J Plant Physiol 133: 460–465

    Google Scholar 

  • Perales EH and Schieder O (1993) Plant Cell Tiss Org Cult 34: 71–76

    Google Scholar 

  • Predieri S, Fasolo F, Passey AJ, Ridout MS, James DJ (1989) J Hort Sci 64: 553–559

    Google Scholar 

  • Sriskandarajah S, Skirvin RM, Abu-Qaoud H, Korban SS (1990) J Hort Sci 65: 113–121

    Google Scholar 

  • Sriskandarajah S, Goodwin PB, Speirs J (1994) Plant Cell Tiss Org Cult 36: 317–329

    Google Scholar 

  • Vancanneyt G, Schmidt R, O'Connor-Sanchez A, Willmitzer L, Rocha-Sosa M (1990) Mol Gen Genet 220: 245–250

    CAS  PubMed  Google Scholar 

  • Van Nieuwkerk JP, Zimmerman RH, Fordham I (1985) HortSci 20: 523

    Google Scholar 

  • Welander M, Maheswaran G (1992) Plant Physiol 140: 223–228

    Google Scholar 

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Authors and Affiliations

  1. F. A. Janssens Laboratorium voor Genetica, Katholieke Universiteit Leuven, Willem de Croylaan 42, B-3001, Heverlee, Belgium

    An De Bondt, Kristel Eggermont, Iris Penninckx, Inge Goderis & Willem F. Broekaert

  2. N. V. Jo Nicolai & Co, Gorsemdorp 51, B-3803, Saint-Truiden, Belgium

    An De Bondt

Authors
  1. An De Bondt
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  2. Kristel Eggermont
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  3. Iris Penninckx
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  4. Inge Goderis
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  5. Willem F. Broekaert
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Communicated by A. M. Boudet

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De Bondt, A., Eggermont, K., Penninckx, I. et al. Agrobacterium-mediated transformation of apple (Malus x domestica Borkh.): an assessment of factors affecting regeneration of transgenic plants. Plant Cell Reports 15, 549–554 (1996). https://doi.org/10.1007/BF00232992

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  • DOI: https://doi.org/10.1007/BF00232992

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