Abstract
In vitro fertilization system of higher plants has been well established using maize gametes and central cells, which can produce embryos and endosperms. In the present study, procedures for isolating gametes and central cells from rice (Oryza sativa L. cv. Nipponbare), a model plant, are reported with the goal of establishing rice in vitro fertilization system. Egg cells and central cells were isolated by manual manipulation of enzyme-treated unpollinated ovules, and an alternative direct isolation method for egg cells that does not use enzymatic treatment was also established. Fluorescent visualization of the granular structures in the cytoplasm of isolated egg cells and the nucleoli in two polar nuclei of isolated central cells suggest that these cells are reliable gametes and central cells. For sperm cell isolation, the contents of rice pollen grains were released by osmotic pressure-induced bursting of the grains. In addition, electrofusion with isolated gametes was successfully conducted.
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Abbreviations
- EDTA:
-
Ethylenediaminetetraacetic acid
- FDA:
-
Fluorescein diacetate
- PMSF:
-
Phenylmethylsulfonyl fluoride
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Acknowledgements
This work was supported in part by Grants-in-Aid from the Ministry of Education, Science, Sport, and Culture of Japan (Grant nos. 16027242 and 17657075 to T.O.) and by Selective Research Fund from Tokyo Metropolitan University to T.O. (Heisei 17 fiscal year).
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Uchiumi, T., Komatsu, S., Koshiba, T. et al. Isolation of gametes and central cells from Oryza sativa L.. Sex Plant Reprod 19, 37–45 (2006). https://doi.org/10.1007/s00497-006-0020-x
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DOI: https://doi.org/10.1007/s00497-006-0020-x