A large-scale Agrobacterium-mediated transformation procedure with a strong positive-negative selection for gene targeting in rice (Oryza sativa L.).
A large-scale transformation procedure handling an adequate number of stable transformants with highly efficient positive-negative selection is a necessary prerequisite to successful gene targeting by homologous recombination, as the integration of a transgene by somatic homologous recombination in higher plants has been reported to be 10(-3) to 10(-5) compared with random integration by non-homologous end joining. We established an efficient and large-scale Agrobacterium-mediated rice transformation protocol that generated around 10(3) stable transformants routinely from 150 seeds and a strong positive-negative selection procedure that resulted in survivors at 10(-2) using the gene for diphtheria toxin A fragment as a negative marker. The established transformation procedure provides a basis for efficient gene targeting in rice.
Terada
R
R
National Institute for Basic Biology, 444-8585, Okazaki, Japan.
Asao
H
H
Iida
S
S
D004167:Diphtheria Toxin / Q000235:genetics; D018390:Gene Targeting; D005819:Genetic Markers; D005822:Genetic Vectors; D012275:Oryza / Q000235:genetics*; D010446:Peptide Fragments / Q000235:genetics; D030821:Plants, Genetically Modified / Q000235:genetics*; D011995:Recombination, Genetic; D012231:Rhizobium / Q000235:genetics*; D014170:Transformation, Genetic
D004167:Diphtheria Toxin; D005819:Genetic Markers; D010446:Peptide Fragments; C029795:diphtheria toxin fragment A