Plant Cell Reports

, 28:387

High-efficiency Agrobacterium-mediated transformation of citrus via sonication and vacuum infiltration

  • Maria Luiza Peixoto de Oliveira
  • Vicente J. Febres
  • Marcio Gilberto Cardoso Costa
  • Gloria A. Moore
  • Wagner Campos Otoni
Genetic Transformation and Hybridization

Abstract

An improved method for the Agrobacterium infiltration of epicotyl segments of ‘Pineapple’ sweet orange [Citrus sinensis (L.) Osbeck] and ‘Swingle’ citrumelo [Citrus paradisi Macf. X Poncirus trifoliata (L.) Raf.] was developed in order to increase transformation frequency. Sonication-assisted Agrobacterium-mediated transformation (SAAT), vacuum infiltration, and a combination of the two procedures were compared with conventional Agrobacterium-mediated inoculation method (‘dipping’ method). It was observed that the morphogenic potential of the epicotyl segments decreased as the duration of SAAT and vacuum treatments increased. Transient GUS expression was not affected by the different SAAT treatments, but it was significantly enhanced by the vacuum infiltration treatments. The highest transformation efficiencies were obtained when the explants were subjected to a combination of SAAT for 2 s followed by 10 min of vacuum infiltration. PCR and Southern blot analysis of the uidA gene were used to confirm the integration of the transgenes. The transformation frequencies achieved in this study (8.4% for ‘Pineapple’ sweet orange and 11.2% for ‘Swingle’ citrumelo) are the highest ones reported for both cultivars.

Keywords

Agrobacterium tumefaciens Orange SAAT Agroinfiltration 

Abbreviations

BAP

6-Benzylaminopurine

NAA

Naphtaleneacetic acid

GUS

β-Glucuronidase

nptII

Neomycin phosphotransferase II

SAAT

Sonication-assisted Agrobacterium-mediated transformation

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Copyright information

© Springer-Verlag 2008

Authors and Affiliations

  • Maria Luiza Peixoto de Oliveira
    • 1
  • Vicente J. Febres
    • 2
  • Marcio Gilberto Cardoso Costa
    • 3
  • Gloria A. Moore
    • 2
  • Wagner Campos Otoni
    • 1
  1. 1.Plant Biology Department, Plant Tissue Culture Laboratory/BIOAGROFederal University of ViçosaViçosaBrazil
  2. 2.Horticultural Sciences Department, Institute of Food and Agricultural Sciences, Plant Molecular and Cellular Biology ProgramUniversity of FloridaGainesvilleUSA
  3. 3.Biological Sciences DepartmentState University of Santa Cruz (UESC)IlhéusBrazil

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