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Highly efficient direct seed transformation protocol for japonica rice (Oryza sativa L.) by Agrobacterium tumefaciens
* 1, 2 , 1, 3
1  Graduate School of Science, Nagoya City University, Mizuho-cho, Mizuho-ku Nagoya, Japan.
2  Bangladesh Institute of Research and Training on Applied Nutrition (BIRTAN), Narayanganj, Bangladesh
3  Research Center for Biological Diversity, Nagoya City University, Mizuho-cho, Mizuho-ku Nagoya, Japan.
Academic Editor: Rodomiro Ortiz

Abstract:

Molecular breeding and gene function studies in plants require high transformation efficiency. Agrobacterium-mediated transformation has contributed significantly to molecular research in many plants, but is inefficient and inconsistent in rice that do not host Agrobacterium. Transformation efficiency in rice remains low. Therefore, this study aimed to establish a simple and efficient transformation method for rice using Agrobacterium. Two foreign genes (CISP1-GFP and CISP2-GFP) and a Agrobacterium strain (EHA105) were used in the experiments. Then, Agrobacterium infection of rice seeds that had absorbed water and germinated under reduced pressure infiltration conditions showed that an average of 14% of the seeds formed after growth (12% with CISP1-GFP and 16% with CISP2-GFP) carried the foreign gene, and it was also confirmed by PCR, Western blot, GFP fluorescence and TAIL-PCR. Since this method does not involve callus formation or re-differentiation of rice plants, no special equipment or complicated operations are required, and transformants can be obtained in only three months. Therefore, this method is expected to simplify rice genetic manipulation and promote molecular breeding of rice.

Keywords: Protocol, direct seed transformation, Western blot, TAIL-PCR, Agrobacterium and Rice (Oryza sativa L.)

 
 
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