Abstract
Grain hardness of wheat is determined by the hardness (Ha)-locus region, which contains three friabilin-related genes: puroindoline-a (Pina), puroindoline-b (Pinb) and GSP-1. In our previous study, we produced the transgenic rice plants harboring the large genomic fragment of the Ha-locus region of Aegilops tauschii containing Pina and GSP-1 genes by Agrobacterium-mediated transformation. To examine the effects of the transgenes in the rice endosperms, we firstly confirmed the homozygosity of the T-DNAs in four independent T2 lines by using fluorescence in situ hybridization (FISH) and DNA gel blot analyses. The transgenes, Pina and GSP-1, were stably expressed in endosperms of the T3 and T4 seeds at RNA and protein levels, indicating that the promoters and other regulatory elements on the wheat Ha-locus region function in rice, and that multigene transformation using a large genomic fragment is a useful strategy. The functional contribution of the transgene-derived friabilins to the rice endosperm structure was considered as an increase of spaces between compound starch granules, resulting in a high proportion of white turbidity seeds.
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Abbreviations
- FISH:
-
Fluorescence in situ hybridization
- GSP-1 :
-
Grain Softness Protein-1
- Ha :
-
Hardness
- Pina :
-
Puroindoline-a
- Pinb :
-
Puroindoline-b
- RT-PCR:
-
Reverse transcription polymerase chain reaction
- SEM:
-
Scanning electron microscope
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Acknowledgments
The authors are highly obliged to Drs. Yasunori Nakamura and Akiko Kubo of the Akita Prefectural University, Dr. Tatsuya M. Ikeda of the National Agricultural Research Center for Western Region, and Dr. Noriaki Aoki of the National Institute of Crop Science for their useful technical and scientific advice. This work was supported in part by the Ministry of Education, Culture, Sports, Science, and Technology of Japan (MEXT) and the Japan Society for the Promotion of Science (JSPS) grants, KAKENHI (18075003 and 20780240 to G.S., 22580004 and 19380194 to Y.M.).
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Communicated by K. Toriyama.
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Online Resource 1: Separated total DNAs (HindIII digested) stained with ethidium bromide before Southern blotting (Fig. 2a) as the DNA amount control (DOC 189 kb)
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Online Resource 2 The CBB-stained SDS-PAGE gels for the protein amount control of Western blot analysis (Fig. 4). (DOC 746 kb)
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Suzuki, G., Wada, H., Goto, H. et al. Transgenic rice plants harboring the grain hardness-locus region of Aegilops tauschii . Plant Cell Rep 30, 2293–2301 (2011). https://doi.org/10.1007/s00299-011-1134-7
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DOI: https://doi.org/10.1007/s00299-011-1134-7