Abstract
Conditions for maximising transient expression of GUS in leaf mesophyll protoplasts of Arabidopsis thaliana ecotype C24 were investigated. It was found that the factors most influencing expression levels, with optimum levels in parenthesis, were plasmid DNA quantity (100 μg per 5 × 105 protoplasts), inclusion of carrier DNA (50 μg), PEG pH and amount (pH above 6, and total PEG concentration at least 9% w/w) and the topological form of the DNA. Linearised plasmid DNA with long flanking sequences 3′ and 5′ to the marker gene yielded the highest levels of GUS expression.
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Abbreviations
- 2,4-d :
-
2,4-dichlorophenoxyacetic acid
- GUS:
-
β-glucuronidase
- MU:
-
methylumbelliferone
- PEG:
-
polyethylene glycol
- X-gluc:
-
5-bromo-4-chloro-3-indolyl-β-glucuronic acid
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Hoffman, A., Halfter, U. & Morris, PC. Transient expression in leaf mesophyll protoplasts of Arabidopsis thaliana . Plant Cell Tiss Organ Cult 36, 53–58 (1994). https://doi.org/10.1007/BF00048315
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DOI: https://doi.org/10.1007/BF00048315