An improved method for protoplast micro-injection suitable for transfer of entire plant chromosomes |
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| Institution: | 1. Tasmanian Institute for Agriculture, University of Tasmania, Hobart, TAS 7001, Australia;2. Collaborative Innovation Centre for Grain Industry, College of Agriculture, Yangtze University, Jingzhou 434025, China;3. College of Agriculture and Biotechnology, Zhejiang University, Hangzhou 310058, China;4. School of Science and Health, Western Sydney University, Penrith, NSW 2751, Australia;5. Hawkesbury Institute for the Environment, Western Sydney University, Penrith, NSW 2751, Australia;6. International Research Centre for Environmental Membrane Biology, Foshan University, Foshan 528000, China |
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| Abstract: | A simple protoplast culture system, combining optimal plating efficiency and the prerequisites for microinjection (immobilization, localization, optics) is described. Microinjection of lucifer yellow into protoplasts and protoplast-derived cells from Nicotiana plumbaginifolia cell suspensions was carried out to demonstrate the vitality of the procedure, the possibility to penetrate cell walls, and the occurrence of aberrant cell divisions early during protoplast regeneration at high frequency.Using specific adaptations of the equipment, transfer of particles was carried out: metaphase chromosomes isolated from a partially synchronized kanamycin-resistant suspension culture of N. plumbaginifolia were microinjected into recipient wild-type N. plumbaginifolia protoplasts. So far only visual evidence was achieved; genetic and molecular verification for chromosome-mediated gene transfer is in progress. |
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