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Title: Agrobacterium-mediated transformation of Mexican lime (Citrus aurantifolia Swingle) using optimized systems for epicotyls and cotelydons

Author
item Oliveira, Maria
item MOORE, GLORIA - University Of Florida
item Thomson, James - Jim
item Stover, Eddie

Submitted to: Advances in Bioscience and Biotechnology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/13/2015
Publication Date: 11/16/2015
Citation: De Oliveira, M.L.P, Moore, G., Thomson, J.G., Stover, E.W. 2015. Agrobacterium-mediated transformation of Mexican lime (Citrus aurantifolia Swingle) using optimized systems for epicotyls and cotelydons. Advances in Bioscience and Biotechnology. 6:657-668.

Interpretive Summary: Several protocols for regeneration of transgenic citrus plants have been reported in the past few decades, but some varieties remain recalcitrant to transformation. This study describe an alternative and reliable protocol for Agrobacterium-mediated transformation and shoot regeneration of citrus from cotyledons after evaluation of the effects of different parameters on organogenesis and transformation efficiencies. To our knowledge, this is the first time that cotyledons have been used as explants for regeneration of transgenic citrus plants through a shoot organogenesis pathway.

Technical Abstract: Epicotyl and internodal stem segments provide the predominantly used explants for regeneration of transgenic citrus plants following co-cultivation with Agrobacterium. Previous reports using epicotyls segments from Mexican lime have shown low affinity for Agrobacterium tumefaciens infection which results in low transformation efficiency. Therefore we decided to explore the use of cotyledons as an explant source. In the present study we examined the timing and frequency of transformation between the conventional protocol, using epicotyls as explants, and an alternative optimized protocol for Agrobacterium-mediated transformation in Mexican lime. The alternative protocol using cotyledons from mature seeds provided an improvement in regeneration capacity, longer shoots and robust rooting, with a very simple transformation procedure, reducing the timing for transgenic Mexican lime plant-recovery. Application of this procedure may also accelerate efficient recovery of others citrus recalcitrant genotypes from in vitro transformation.