Controlled cell wall regeneration for efficient microinjections of Nicotiana tabacum var. Carlson protoplasts |
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Authors: | S Joshi A M Vincentini |
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Institution: | (1) Department of Microbiology, University of Toronto, 100 College Street, #214, M5G 1L5 Toronto, Ontario, Canada |
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Abstract: | Nicotiana tabacum var. Carlson protoplast culture conditions were modified to contain a cell wall inhibitor, 2,6-dichlorobenzonitrile, in order to delay cell wall regeneration and to allow efficient nuclear and cytoplasmic microinjections. Under modified conditions, the protoplast preparations appeared healthier as compared to the control protoplasts and showed no resistance at all during microinjection. Furthermore, the duration of protoplast microinjection was extended for up to 3–4 days. In order to set up nuclear microinjections, the nuclei of these protoplasts were stained either before or after immobilization without any adverse effect on their mitotic activity. Successful cytoplasmic microinjections were demonstrated by injecting Alfalfa mosaic virus (AMV) RNA, which resulted in viral infection of 14% of the injected protoplasts.Abbreviations AMV
Alfalfa Mosaic virus
- BAP
6-benzylaminopurine
- 2,4-D
2,4-dichlorophenoxy-acetic acid
- DB
2,6-dichlorobenzonitrile
- LR
lissamine rhodamine
- NAA
1-naphthalene-acetic acid |
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Keywords: | |
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