Plant Regeneration and Its Functional Analysis within Transgenic Rice of ASG-1, an Apomixis-specific Gene Isolated from Apomictic Guinea Grass
Lanzhuang Chen *
Graduated School of Horticultural and Food Science, Minami Kyushu University, 3764-1, Tatenocho, Miyakonojo City, Miyazaki, 885-0035, Japan
Liming Guan
Faculty of Education and Culture, University of Miyazaki, 1-1, Gakuen Kibanadai, Miyazaki, 889-2192, Japan
Daiki Toyomoto
Graduated School of Horticultural and Food Science, Minami Kyushu University, 3764-1, Tatenocho, Miyakonojo City, Miyazaki, 885-0035, Japan
Toru Sugita
Miyazaki Prefectural Agricultural Station, 5805, Shimonaka, Sadowara, Miyazaki, 880-0212, Japan
Takuro Hamaguchi
Miyazaki Prefecture Office, 2-10-1, Tachibanatori-Higashi, Miyazaki, 880-8501, Japan
Reiji Okabe
Miyazaki Prefecture Office, 2-10-1, Tachibanatori-Higashi, Miyazaki, 880-8501, Japan
*Author to whom correspondence should be addressed.
Abstract
Aims: To analyze the function of the ASG-1, an apomixis-specific gene has been isolated from apomictic guinea grass, the combination of Agrobacterium-mediated transformation system and pSMA35H2-NG binary vector were used for transformation of ASG-1 to ‘Nipponbare’ of rice, and its functional analysis of transgenic rice was carried out.
Study Design: 1) plant regeneration of transgenic rice of ASG-1 was achieved, and 2) it was followed by experiments with the obtained non-graining ear in the whole plants (NGEP) using comparative observation in both pollens of guinea grass and the T0 NGEP with Nomarski differential interference-contrast microscopy (DIC).
Place and Duration of Study: Faculty of Environmental and Horticultural Science, Minami Kyushu University, between September 2012 and October 2015.
Methodology: As a preliminary transformation, a β-glucuronidase (GUS) was used to infect the callus. Plant regeneration of the transgenic rice of ASG-1 were achieved. DNAs of the transgenic rice were used for detection of ASG-1 by PCR. For analysis of NGEP, the anthers and ovaries of the transgenic rice and guinea grass were used for comparative observation with DIC.
Results: 1) GUS was successfully expressed in calli of both rice and guinea grass. 2) The transgenic rice plants of ASG-1 were then obtained from the culture of seed-derived calli. 3) The natures of T0 callus and T1 plants were confirmed. 4) A subset (~15%) of T0 plants showed morphological sterility with NGEP. 5) The ovaries of the NGEP showed identical characteristics to “Nipponbare” and other normal T1 plants; however, the pollen showed normal rates of <38%. 6) The pollen of guinea grass at anthesis showed rates of normal pollen as low as that of the NGEP rice.
Conclusion: ASG-1 may play a role in regulating pollen sterility. Comparing these findings with our previous reports, we identified co-localization for ASG-1 in pollen of both rice and guinea grass, suggesting that the trait of ASG-1 influencing pollen sterility might be inherited from guinea grass to rice through the transformation process.
Keywords: Apomixis, ASG-1 (apomixis-specific gene 1), guinea grass (Panicum maximum Jacq.), NGEP (non-graining ear formation in whole plant), pollen sterility, transgenic rice (Oryza sativa L.)