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Plant Cell Tissue Organ Cult. 2009;96(3):325-333.

Development of an Agrobacterium-mediated transformation protocol for the tree-legume Leucaena leucocephala using immature zygotic embryos.

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1
Department of Molecular Biosciences and Bioengineering, University of Hawaii at Manoa, 1955 East-West Road, Honolulu 96822, HI, USA.

Abstract

The tree-legume Leucaena leucocephala (leucaena) is used as a perennial fodder because of its fast-growing foliage, which is high in protein content. The use of leucaena as a fodder is however restricted due to the presence of the toxin mimosine. Improvements in the nutritional contents as well as other agronomic traits of leucaena can be accomplished through genetic transformation. The objective of this research was to develop a transformation protocol for leucaena using phosphinothricin resistance as the plant selectable marker. Explants obtained from immature zygotic embryos infected with the Agrobacterium tumefaciens strain C58C1 containing the binary plasmid pCAMBIA3201 produced four putative transformed leucaena plants. Transformation was con- firmed by PCR, RT-PCR, Southern blot, Western analyses, GUS-specific enzyme activity and herbicide leaf spraying assay. A transformation efficiency of 2% was established using this protocol.

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