Influence of enzyme solution on protoplast culture and transient gene expression in maize (Zea mays L.) |
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Authors: | Brigitte Krautwig Paul A. Lazzeri Horst Lörz |
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Affiliation: | (1) Institute of General Botany, AMP II, University of Hamburg, Ohnhorst-strasse 18, 22609 Hamburg, Germany;(2) Biochemistry and Physiology Department, Institute of Arable Crops Research, Rothamsted Experimental Station, AL5 2JQ Harpenden, Hertfordshire, UK |
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Abstract: | The influence of two enzyme solutions, differing only in the presence or absence of Macerozyme, on protoplast yield, colony formation and transient GUS (-glucuronidase) activity was studied. For all parameters tested the presence of Macerozyme during protoplast isolation had a negative influence. Using an enzyme solution without Macerozyme suspension aggregates gave up to 4.4 times higher protoplast yield and plating efficiencies were increased up to 10-fold. Further, protoplasts isolated without macerozyme showed a 5.2-fold higher GUS activity in transient gene expression. Apart from the presence of Macerozyme, longer incubation (3 compared with 1.5 h) of cell aggregates in the enzyme solution also had a negative effect on transient transformation efficiency. These data demonstrate that protoplast isolation conditions have a profound effect on transient gene expression and it is proposed that these factors will also influence stable transformation efficiency.Abbreviations CP cellulase pectolyase - CPM cellulase pectolyase Macerozyme - 2,4-d 2,4-dichlorophenoxyacetic acid |
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Keywords: | /content/x6450544w05t57j7/xxlarge946.gif" alt=" beta" align=" MIDDLE" BORDER=" 0" >-glucuronidase Macerozyme plating efficiency protoplast yield |
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