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High frequency Agrobacterium tumefaciens-mediated plant transformation induced by ammonium nitrate

  • Genetic Transformation and Hybridization
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Abstract

Success in plant genetic transformation depends on the efficiency of explant regeneration and transgene integration. Whereas the former one depends on explant totipotency, the latter depends on the activity of host DNA repair and chromatin organisation factors. We analyzed whether factors that result in an increase in recombination frequency can also increase transformation efficiency. Here, we report that a threefold increase in the concentration of NH4NO3 in the growth medium results in more than a threefold increase in the Agrobacterium tumefaciens-mediated transformation frequency of Nicotiana tabacum plants. Regeneration of calli without selection showed that the increase in transformation frequency was primarily due to the increase in transgene integration efficiency rather than in tissue regeneration efficiency. PCR analysis of insertion sites showed a decrease in the frequency of truncations of the T-DNA right border and an increase on the left border. We hypothesize that exposure to ammonium nitrate modifies the activity of host factors leading to higher frequency of transgene integrations and possibly to the shift in the mechanism of transgene integrations.

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Abbreviations

HR:

Homologous recombination

NHEJ:

Non-homologous end-joining

DSB:

Double strand break

ssDNA:

Single stranded DNA

ssT-DNA:

Single stranded T-DNA

dsT-DNA:

Double stranded T-DNA

HRF:

Homologous recombination frequency

RR:

Recombination rate

CRE:

Callus regeneration efficiency

STF:

Stable transformation frequency

GUS:

uidA (β-glucuronidase) gene

LUC:

Luciferase gene

DSBR:

DSB repair (model)

SSGR:

Single-strand-gap repair (model)

Dpg:

Days post germination

MS:

Murashige-Skoog (media)

References

  • Alonso JM, Stepanova AN, Leisse TJ, Kim CJ, Chen H, Shinn P, Stevenson DK, Zimmerman J, Barajas P, Cheuk R, Gadrinab C, Heller C, Jeske A, Koesema E, Meyers CC, Parker H, Prednis L, Ansari Y, Choy N, Deen H, Geralt M, Hazari N, Hom E, Karnes M, Mulholland C, Ndubaku R, Schmidt I, Guzman P, Aguilar-Henonin L, Schmid M, Weigel D, Carter DE, Marchand T, Risseeuw E, Brogden D, Zeko A, Crosby WL, Berry CC, Ecker JR (2003) Genome-wide insertional mutagenesis of Arabidopsis thaliana. Science 301:653–657

    Article  PubMed  Google Scholar 

  • Basnakian AG, James SJ (1996) Quantification of 3′OH DNA breaks by random oligonucleotideprimed synthesis (ROPS) assay. DNA Cell Biol 15(3):255–262

    Article  PubMed  CAS  Google Scholar 

  • Bezzubova O, Silbergleit A, Yamaguchi-Iwai Y, Takeda S, Buerstedde JM (1997) Reduced X-ray resistance and homologous recombination frequencies in a RAD54−/− mutant of the chicken DT40 cell line. Cell 89:185–193

    Article  PubMed  CAS  Google Scholar 

  • Bleuyard JY, Gallego ME, White CI (2006) Recent advances in understanding of the DNA double-strand break repair machinery of plants. DNA Repair (Amst) 5(1):1–12

    Article  CAS  Google Scholar 

  • Boyko A, Filkowski J, Hudson D, Kovalchuk I (2006) Homologous recombination in plants is organ specific. Mutat Res 595(1–2):145–155

    PubMed  CAS  Google Scholar 

  • Boyko A, Kathiria P, Zemp FJ, Yao Y, Pogribny I, Kovalchuk I (2007) Transgenerational changes in the genome stability and methylation in pathogen-infected plants: (virus-induced plant genome instability). Nucleic Acids Res 35(5):1714–1725

    Article  PubMed  CAS  Google Scholar 

  • Brunaud V, Balzergue S, Dubreucq B, Aubourg S, Samson F, Chauvin S, Bechtold N, Cruaud C, DeRose R, Pelletier G, Lepiniec L, Caboche M, Lecharny A (2002) T-DNA integration into the Arabidopsis genome depends on sequences of pre-insertion sites. EMBO Rep 3(12):1152–1157

    Article  PubMed  CAS  Google Scholar 

  • Chen F, Nastasi A, Shen Z, Brenneman M, Crissman H, Chen DJ (1997) Cell cycle-dependent protein expression of mammalian homologs of yeast DNA double strand break repair genes RAD51 and RAD52. Mutat Res 384:205–211

    PubMed  CAS  Google Scholar 

  • Chilton M-DM, Que Q (2003) Targeted integration of T-DNA into the tobacco genome at double-strand breaks: new insights on the mechanism of T-DNA integration. Plant Physiol 133:956–965

    Article  PubMed  CAS  Google Scholar 

  • Choi Y-E, Yang D-C, Choi K-T (1998) Induction of somatic embryos by macrosalt stress from mature zygotic embryos of Panax ginseng. Plant Cell Tissue Cult Organ Cult 52:117–181

    Article  Google Scholar 

  • Chugh A, Khurana P (2002) Gene expression during somatic embryogenesis—recent advances. Curr Sci 83(6):715–729

    CAS  Google Scholar 

  • Citovsky V, Kozlovsky SV, Lacroix B, Zaltsman A, Dafny-Yelin M, Vyas S, Tovkach A, Tzfira T (2007) Biological systems of the host cell involved in Agrobacterium infection. Cell Microbiol 9(1):9–20

    Article  PubMed  CAS  Google Scholar 

  • Cline MG, Thangavelu M, Dong ILK (2006) A possible role of cytokinin in mediating long-distance nitrogen signalling in the promotion of sylleptic branching in hybrid poplar. J Plant Physiol 163(6):684–688

    Article  PubMed  CAS  Google Scholar 

  • Cottage A, Yang A, Maunders H, de Lacy RC, Ramsay NA (2001) Identification of DNA sequences flanking T-DNA insertions by PCR-walking. Plant Mol Biol Reporter 19:321–327

    Article  CAS  Google Scholar 

  • Crawford NM (1995) Nitrate: nutrient and signal for plant growth. Plant Cell 7(7):859–868

    Article  PubMed  CAS  Google Scholar 

  • De Block M, Debrouwer D, Moens T (1997) The development of a nuclear male sterility system in wheat. Expression of the barnase gene under the control of tapetum specific promoters. Theor Appl Genet 95:125–131

    Article  Google Scholar 

  • Dias JS, Martins MG (1999) Effect of silver nitrate on anther culture embryo production of different Brassica oleracea morphotypes. Sci Hortic 82:299–307

    Article  CAS  Google Scholar 

  • Dudas A, Chovanec M (2004) DNA double-strand break repair by homologous recombination. Mutat Res 566:131–167

    Article  PubMed  CAS  Google Scholar 

  • Endo M, Ishikawa Y, Osakabe K, Nakayama S, Kaya H, Araki T, Shibahara K, Abe K, Ichikawa H, Valentine L, Hohn B, Toki S (2006) Increased frequency of homologous recombination and T-DNA integration in Arabidopsis CAF-1 mutants. Europ Mol Biol Org J 25(23):5579–5590

    CAS  Google Scholar 

  • Essers J, Hendriks RW, Swagemakers SM, Troelstra C, de Wit J, Bootsma D, Hoeijmakers JH, Kanaar R (1997) Disruption of mouse RAD54 reduces ionizing radiation resistance and homologous recombination. Cell 89:195–204

    Article  PubMed  CAS  Google Scholar 

  • Fladung M (1999) Gene stability in transgenic aspen (Populus). I. Flanking DNA sequences and T-DNA structure. Mol Gen Genet 260(6):574–581

    Article  PubMed  CAS  Google Scholar 

  • Forde BG (2002a) Local and long-range signalling pathways regulating plant responses to nitrate. Annu Rev Plant Biol 53:203–224

    Article  PubMed  CAS  Google Scholar 

  • Forde BG (2002b) The role of long-distance signalling in plant responses to nitrate and other nutrients. J Exp Bot 53(366):39–43

    Article  PubMed  CAS  Google Scholar 

  • Francis KE, Spiker S (2005) Identification of Arabidopsis thaliana transformants without selection reveals a high occurrence of silenced T-DNA integrations. Plant J 41(3):464–477

    Article  PubMed  CAS  Google Scholar 

  • Gherbi H, Gallego ME, Jalut N, Lucht JM, Hohn B, White CI (2001) Homologous recombination in planta is stimulated in the absence of Rad50. Europ Mol Biol Org Rep 2(4):287–291

    CAS  Google Scholar 

  • Gorbunova V, Levy AA (1999) How plants make ends meet: DNA double strand break repair. Trends Plant Sci 4:263–269

    Article  PubMed  Google Scholar 

  • Gordon-Kamm W, Dilkes BP, Lowe K, Hoerster G, Sun X, Ross M, Church L, Bunde C, Farrell J, Hill P, Maddock S, Snyder J, Sykes L, Li Z, Woo YM, Bidney D, Larkins BA (2002) Stimulation of the cell cycle and maize transformation by disruption of the plant retinoblastoma pathway. Proc Natl Acad Sci USA 99(18):11975–11980

    Article  PubMed  CAS  Google Scholar 

  • Grimes HD, Hodges TK (1990) The inorganic NO3 −:NH4 + ratio influences plant regeneration and auxin sensitivity in primary callus derived from immature embryos of indica rice (Oryza sativa L.). J Plant Physiol 136:362–367

    CAS  Google Scholar 

  • Halperin W (1966) Alternative morphogenetic events in cell suspensions. Am J Bot 53:443–453

    Article  Google Scholar 

  • Hanin M, Paszkowski J (2003) Plant genome modification by homologous recombination. Curr Opin Plant Biol 6(2):157–162

    Article  PubMed  CAS  Google Scholar 

  • Hanin M, Mengiste T, Bogucki A, Paszkowski J (2000) Elevated levels of intrachromosomal homologous recombination in Arabidopsis overexpressing the MIM gene. Plant J 24(2):183–189

    Article  PubMed  CAS  Google Scholar 

  • He D-G, Yang Y-M, Scott KJ (1989) The effect of macroelements in the induction of embryogenic callus from immature embryos of wheat (Triticum aestivum L.). Plant Sci 64:251–258

    Article  CAS  Google Scholar 

  • He D-G, Yang Y-M, Scott KJ (1991) Zinc deficiency and the formation of white structures in immature embryo cultures of wheat (Triticum aestivum L.). Plant Cell Tissue Organ Cult 24(1):9–12

    Article  CAS  Google Scholar 

  • Ilnytskyy Y, Boyko A, Kovalchuk I (2004) Luciferase-based transgenic recombination assay is more sensitive than beta-glucoronidase-based. Mutat Res 559(1–2):189–197

    PubMed  CAS  Google Scholar 

  • Immonen AST (1996) Influence of media and growth regulators on somatic embryogenesis and plant regeneration for production of primary triticales. Plant Cell Tissue Organ Cult 44(1):45–52

    Article  CAS  Google Scholar 

  • Jiménez JM (2001) Regulation of in vitro somatic embryogenesis with emphasis on the role of endogenous hormones. Revista Brasileira de Fisiologia Vegetal 13(2):196–223

    Article  Google Scholar 

  • Khanna HK, Daggard GE (2003) Agrobacterium tumefaciens-mediated transformation of wheat using a superbinary vector and a polyamine-supplemented regeneration medium. Plant Cell Rep 21(5):429–436

    PubMed  CAS  Google Scholar 

  • Kim S-I, Veena, Gelvin SB (2007) Genome-wide analysis of Agrobacterium T-DNA integration sites in the Arabidopsis genome generated under non-selective conditions. Plant J 51:779–791

  • Kothari SL, Agarwal K, Kumar S (2004) Inorganic nutrient manipulation for highly improved in vitro plant regeneration in finger millet—Elusine coracana (L.) gaertn. In Vitro Cell Dev Biol Plant 40:515–519

    Article  CAS  Google Scholar 

  • Kovalchuk I, Kovalchuk O, Hohn B (2000) Genome-wide variation of the somatic mutation frequency in transgenic plants. EMBO J 19(17):4431–4438

    Article  PubMed  CAS  Google Scholar 

  • Kumar S, Fladung M (2002) Transgene integration in aspen: structures of integration sites and mechanism of T-DNA integration. Plant J 31(4):543–551

    Article  PubMed  CAS  Google Scholar 

  • Li DD, Shi W, Deng XX (2003) Factors influencing Agrobacterium-mediated embryogenic callus transformation of Valencia sweet orange (Citrus sinensis) containing the pTA29-barnase gene. Tree Physiol 23(17):1209–1215

    PubMed  CAS  Google Scholar 

  • Li J, Vaidya M, White C, Vainstein A, Citovsky V, Tzfira T (2005) Involvement of KU80 in T-DNA integration in plant cells. Proc Natl Acad Sci USA 102(52):19231–19236

    Article  PubMed  CAS  Google Scholar 

  • Lida S, Terada R (2004) A tale of two integrations, transgene and T-DNA: gene targeting by homologous recombination in rice. Curr Opin Biotechnol 15:132–138

    Article  CAS  Google Scholar 

  • Liu Y, Stasiak AZ, Masson J-Y, Mcllwaith MJ, Stasiak A, West SC (2004) Conformational changes modulate the activity of human RAD51 protein. J Mol Biol 337:817–827

    Article  PubMed  CAS  Google Scholar 

  • Maës OC, Chibbar RN, Caswell K, Leung N, Kartha KK (1996) Somatic embryogenesis from isolated scutella of wheat: effects of physical, physiological and genetic factors. Plant Sci 121:75–84

    Article  Google Scholar 

  • Malabadi RB, Staden JV (2006) Cold enhanced somatic embryogenesis in Pinus patula is mediated by calcium. S Afr J Bot 72(4):613–618

    Article  CAS  Google Scholar 

  • Malik MR, Wang F, Dirpaul JM, Zhou N, Hammerlindl J, Keller W, Abrams SR, Ferrie AM, Krochko JE (2008) Isolation of an embryogenic line from non-embryogenic Brassica napus cv. Westar through microspore embryogenesis. J Exp Bot 59:2857–2873

    Article  PubMed  CAS  Google Scholar 

  • Martino-Catt SJ, Sachs ES (2008) Editor’s choice series: the next generation of biotech crops. Plant Physiol 147:3–5

    Article  PubMed  CAS  Google Scholar 

  • Menke-Milczarek I, Zimny J (2001) NH4 + and NO3 − requirements for wheat somatic embryogenesis. Acta Physiol Plant 23(1):37–42

    Article  CAS  Google Scholar 

  • Meyer P, Walgenbach E, Bussmann K, Hombrecher G, Saedler H (1985) Synchronized tobacco protoplasts are efficiently transformed by DNA. Mol Gen Genet 201:513–518

    Article  CAS  Google Scholar 

  • Meza TJ, Stangeland B, Mercy IS, Skårn M, Nymoen DA, Berg A, Butenko MA, Håkelien AM, Haslekås C, Meza-Zepeda LA, Aalen RB (2002) Analyses of single-copy Arabidopsis T-DNA-transformed lines show that the presence of vector backbone sequences, short inverted repeats and DNA methylation is not sufficient or necessary for the induction of transgene silencing. Nucleic Acids Res 30(20):4556–4566

    Article  PubMed  CAS  Google Scholar 

  • Miller AJ, Fan X, Orsel M, Smith SJ, Wells DM (2007) Nitrate transport and signalling. J Exp Bot 58(9):2297–2306

    Article  PubMed  CAS  Google Scholar 

  • Mordhorst AP, Lorz H (1993) Embryogenesis and development of isolated barley (Hordeum vulgare L.) microspores are influenced by the amount and composition of nitrogen sources in culture media. J Plant Physiol 142:485–492

    CAS  Google Scholar 

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

    Article  CAS  Google Scholar 

  • Mysore KS, Nam J, Gelvin SB (2000) An Arabidopsis histone H2A mutant is deficient in Agrobacterium T-DNA integration. Proc Natl Acad Sci USA 97:948–953

    Article  PubMed  CAS  Google Scholar 

  • Ninomiya Y, Suzuki K, Ishii C, Inoue H (2004) Highly efficient gene replacements in Neurospora strains deficient for nonhomologous end-joining. Proc Natl Acad Sci USA 101(33):12248–12253

    Article  PubMed  CAS  Google Scholar 

  • Okada K, Takebe I, Nagata T (1986) Expression and integration of genes into highly synchronized plant protoplasts. Mol Gen Genet 205:398–403

    Article  CAS  Google Scholar 

  • Orel N, Kyryk A, Puchta H (2003) Different pathways of homologous recombination are used for the repair of double-strand breaks within tandemly arranged sequences in the plant genome. Plant J 35:604–612

    Article  PubMed  CAS  Google Scholar 

  • Perl A, Kless H, Blumenthal A, Galili G, Galun E (1992) Improvement of plant regeneration and GUS expression in scutellar wheat calli by optimization of culture conditions and DNA-microprojectile delivery procedures. Mol Gen Genet 235(2–3):279–284

    Article  PubMed  CAS  Google Scholar 

  • Puchta H (2002) Gene replacement by homologous recombination in plants. Trends Plant Sci 1:340–348

    Google Scholar 

  • Puchta H (2005) The repair of double-strand breaks in plants: mechanisms and consequences for genome evolution. J Exp Bot 56:1–14

    Article  PubMed  CAS  Google Scholar 

  • Racusen RH, Schiavone FM (1990) Positional cues and differential gene expression in somatic embryos of higher plants. Cell Differ Dev 30:159–169

    Article  PubMed  CAS  Google Scholar 

  • Reiss B (2003) Homologous recombination and gene targeting in plant cells. Int Rev Cytol 228:85–139

    Article  PubMed  CAS  Google Scholar 

  • Reiss B, Schubert I, Kopchen K, Wendeler E, Schell J, Puchta H (2000) RecA stimulates sister chromatid exchange and the fidelity of double-strand break repair, but not gene targeting, in plants transformed by Agrobacterium. Proc Natl Acad Sci USA 97:3358–3363

    Article  PubMed  CAS  Google Scholar 

  • Rijven AHGC (1958) Effect of some inorganic nitrogenous substances on growth and nitrogen assimilation of young plant embryos. Aust J Biol Sci 11:142–154

    CAS  Google Scholar 

  • Ruffel S, Freixes S, Balzergue S, Tillard P, Jeudy C, Martin-Magniette ML, van der Merwe MJ, Kakar K, Gouzy J, Fernie AR, Udvardi M, Salon C, Gojon A, Lepetit M (2008) Systemic signaling of the plant N status triggers specific transcriptome responses depending on the N source in Medicago truncatula. Plant Physiol 146(4):2020–2035

    Article  PubMed  CAS  Google Scholar 

  • Sahrawat AK, Becker D, Lütticke S, Lörz H (2003) Genetic improvement of wheat via alien gene transfer, an assessment. Plant Sci 165:1147–1168

    Article  CAS  Google Scholar 

  • Salomon S, Puchta H (1998) Capture of genomic and T-DNA sequences during double strand break repair in somatic plant cells. Europ Mol Biol Org J 17:6086–6095

    CAS  Google Scholar 

  • Shaked H, Melamed-Bessudo C, Levy AA (2005) High frequency gene targeting in Arabidopsis plants expressing the yeast RAD54 gene. Proc Natl Acad Sci USA 102:12265–12269

    Article  PubMed  CAS  Google Scholar 

  • Shalev G, Sitrit Y, Avivi-Ragolski N, Lichtenstein C, Levy AA (1999) Stimulation of homologous recombination in plants by expression of the bacterial resolvase ruvC. Proc Natl Acad Sci USA 96:7398–7402

    Article  PubMed  CAS  Google Scholar 

  • Shim KS, Schmutte C, Yoder K, Fishel R (2006) Defining the salt effect on human RAD51 activities. DNA Repair (Amst) 5(6):718–730

    Article  CAS  Google Scholar 

  • Shrivastav M, De Haro LP, Nickoloff JA (2008) Regulation of DNA double-strand break repair pathway choice. Cell Res 18(1):134–147

    Article  PubMed  CAS  Google Scholar 

  • Siebert PD, Chenchik A, Kellogg DE, Lukyanov KA, Lukyanov SA (1995) An improved PCR method for walking in uncloned genomic DNA. Nucleic Acids Res 23:1087–1088

    Article  PubMed  CAS  Google Scholar 

  • Sigurdsson S, Trujillo K, Song BW, Stratton S, Sung P (2001) Basis for avid homologous DNA strand exchange by human Rad 51 and RPA. J Biol Chem 276(12):8798–8806

    Article  PubMed  CAS  Google Scholar 

  • Stitt M (1999) Nitrate regulation of metabolism and growth. Curr Opin Plant Biol 2(3):178–186

    Article  PubMed  CAS  Google Scholar 

  • Swoboda P, Hohn B, Gal S (1993) Somatic homologous recombination in planta: the recombination frequency is dependent on the allelic state of recombining sequences and may be influenced by genomic position effects. Mol Gen Genet 237(1–2):33–40

    PubMed  CAS  Google Scholar 

  • Swoboda P, Gal S, Hohn B, Puchta H (1994) Intrachromosomal homologous recombination in whole plants. Europ Mol Biol Org J 13:484–489

    CAS  Google Scholar 

  • Takei K, Sakakibara H, Taniguchi M, Sugiyama T (2001) Nitrogen-dependent accumulation of cytokinins in root and the translocation to leaf: implication of cytokinin species that induces gene expression of maize response regulator. Plant Cell Physiol 42(1):85–93

    Article  PubMed  CAS  Google Scholar 

  • Tinland B, Schoumacher F, Gloeckler V, Bravo-Angel AM, Hohn B (1995) The Agrobacterium tumefaciens virulence D2 protein is responsible for precise integration of T-DNA into the plant genome. EMBO J 14(14):3585–3595

    PubMed  CAS  Google Scholar 

  • Tzfira T, Frankman R, Vaidya M, Citovsky V (2003) Site-specific integration of Agrobacterium T-DNA via double-stranded intermediates. Plant Physiol 133:1011–1023

    Article  PubMed  CAS  Google Scholar 

  • Tzfira T, Li J, Lacroix B, Citovsky V (2004) Agrobacterium T-DNA integration: molecules and models. Trends Genet 20(8):375–383

    Article  PubMed  CAS  Google Scholar 

  • van Attikum H, Hooykaas PJJ (2003) Genetic requirements for the targeted integration of Agrobacterium T-DNA in Saccharomyces cerevisiae. Nucleic Acids Res 31:826–832

    Article  PubMed  CAS  Google Scholar 

  • Vergunst AC, Hooykaas PJJ (1999) Recombination in the plant genome and its application in biotechnology. Crit Rev Plant Sci 18:1–31

    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 

  • Waldman BC, Waldman AS (1990) Illegitimate and homologous recombination in mammalian cells: differential sensitivity to an inhibitor of poly(ADP-ribosylation). Nucleic Acids Res 18(20):5981–5988

    Article  PubMed  CAS  Google Scholar 

  • Waldman AS, Waldman BC (1991) Stimulation of intrachromosomal homologous recombination in mammalian cells by an inhibitor of poly(ADP-ribosylation). Nucleic Acids Res 19(21):5943–5947

    Article  PubMed  CAS  Google Scholar 

  • Walker KA, Sato SJ (1981) Morphogenesis in callus tissue of Medicago sative: the role of ammonium ion in somatic embryogenesis. Plant Cell Tiss Org Cult 1:109–121

    Article  CAS  Google Scholar 

  • Windels P, De Buck S, Van Bockstaele E, De Loose M, Depicker A (2003) T-DNA integration in Arabidopsis chromosomes. Presence and origin of filler DNA sequences. Plant Physiol 133(4):2061–2068

    Article  PubMed  CAS  Google Scholar 

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Acknowledgments

We would like to thank Alicja Ziemienowicz for critical reading of the manuscript, Elden Van Klei and Viktor Titov for technical assistance with callus regenerations, and Valentina Titova for proofreading the text.

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  1. Department of Biological Sciences, University of Lethbridge, Lethbridge, Canada

    Alex Boyko, Aki Matsuoka & Igor Kovalchuk

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  1. Alex Boyko
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Correspondence to Igor Kovalchuk.

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Communicated by H. Ebinuma.

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Boyko, A., Matsuoka, A. & Kovalchuk, I. High frequency Agrobacterium tumefaciens-mediated plant transformation induced by ammonium nitrate. Plant Cell Rep 28, 737–757 (2009). https://doi.org/10.1007/s00299-009-0676-4

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