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Evaluation of four Agrobacterium tumefaciens strains for the genetic transformation of tomato (Solanum lycopersicum L.) cultivar Micro-Tom

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Abstract

Key message

Agrobacterium tumefaciens strains differ not only in their ability to transform tomato Micro-Tom, but also in the number of transgene copies that the strains integrate in the genome.

Abstract

The transformation efficiency of tomato (Solanum lycopersicum L.) cv. Micro-Tom with Agrobacterium tumefaciens strains AGL1, EHA105, GV3101, and MP90, harboring the plasmid pBI121 was compared. The presence of the nptII and/or uidA transgenes in regenerated T0 plants was determined by PCR, Southern blotting, and/or GUS histochemical analyses. In addition, a rapid and reliable duplex, qPCR TaqMan assay was standardized to estimate transgene copy number. The highest transformation rate (65 %) was obtained with the Agrobacterium strain GV3101, followed by EHA105 (40 %), AGL1 (35 %), and MP90 (15 %). The mortality rate of cotyledons due to Agrobacterium overgrowth was the lowest with the strain GV3101. The Agrobacterium strain EHA105 was more efficient than GV3101 in the transfer of single T-DNA insertions of nptII and uidA transgenes into the tomato genome. Even though Agrobacterium strain MP90 had the lowest transformation rate of 15 %, the qPCR analysis showed that the strain MP90 was the most efficient in the transfer of single transgene insertions, and none of the transgenic plants produced with this strain had more than two insertion events in their genome. The combination of higher transformation efficiency and fewer transgene insertions in plants transformed using EHA105 makes this Agrobacterium strain optimal for functional genomics and biotechnological applications in tomato.

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Abbreviations

MS:

Murashige and Skoog medium

YEP:

Yeast Extract and Peptone medium

Z:

Zeatin

WT:

Wild type

GUS:

β-Glucuronidase

nptII:

Neomycin phosphotransferase

uidA:

β-Glucuronidase

X-Gluc:

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

bp:

Base pair

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Authors and Affiliations

  1. Plant Transformation Research Center, University of California Riverside, CA, Riverside, USA

    V. J. Chetty, D. Garcia, J. Narváez-Vásquez, W. Lopez & M. L. Orozco-Cárdenas

  2. Universidad de Caldas, Manizales, Caldas, Colombia

    N. Ceballos

Authors
  1. V. J. Chetty
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  2. N. Ceballos
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  3. D. Garcia
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  4. J. Narváez-Vásquez
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  5. W. Lopez
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  6. M. L. Orozco-Cárdenas
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Corresponding author

Correspondence to M. L. Orozco-Cárdenas.

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Communicated by B. Li.

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Chetty, V.J., Ceballos, N., Garcia, D. et al. Evaluation of four Agrobacterium tumefaciens strains for the genetic transformation of tomato (Solanum lycopersicum L.) cultivar Micro-Tom. Plant Cell Rep 32, 239–247 (2013). https://doi.org/10.1007/s00299-012-1358-1

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  • DOI: https://doi.org/10.1007/s00299-012-1358-1

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