Transient gene expression in electroporated Picea glauca protoplasts

The reporter gene for chloramphenicol acetyltransferase (CAT) was introduced into white spruce (Picea glauca (Moench) Voss.) protoplasts by electroporation. CAT transient gene expression was increased by increasing the concentration of pCaMVCN plasmid and was affected by the level of the applied vol...

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Bibliographic Details
Published in:Plant cell reports 1988-12, Vol.7 (7), p.481-484
Main Authors: Bekkaoui, F, Pilon, M, Laine, E, Raju, D.S.S, Crosby, W.L, Dunstan, D.I
Format: Article
Language:English
Subjects:
Biological and medical sciences
Biotechnology
chloramphenicol
enzyme activity
Fundamental and applied biological sciences. Psychology
Gene expression
Genetic engineering
Genetic technics
genetic transformation
Methods. Procedures. Technologies
Miscellaneous
Molecular and cellular biology
Molecular genetics
Picea glauca
plasmids
protoplast fusion
Synthetic digonucleotides and genes. Sequencing
transferases
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Summary:The reporter gene for chloramphenicol acetyltransferase (CAT) was introduced into white spruce (Picea glauca (Moench) Voss.) protoplasts by electroporation. CAT transient gene expression was increased by increasing the concentration of pCaMVCN plasmid and was affected by the level of the applied voltage. Highest CAT activities were obtained after electroporation with a pulse of 350V.cm(-1) having an exponential decay constant of approximately 105ms. Linearized plasmid constructs gave much higher levels of CAT activity than circular plasmid. Attempts to use the Escherichia coli β-glucuronidase gene (β-GUS) as a marker gene revealed very high levels of β-GUS-like activity in electroporated protoplasts. This activity was mainly due to a small molecule and may mask successful transformation since β-GUS-like activity increased when plasmid DNA was present during electroporation.
ISSN:0721-7714
1432-203X
DOI:10.1007/BF00272736