Factors influencing transient gene expression in electroporated tall fescue protoplasts |
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Authors: | Ramachandra V Penmetsa and Sam-Bong Ha |
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Institution: | Department of Crop and Soil Environmental Sciences, Virginia Polytechnic Institute and State University, Blacksburg, VA 24061-0404, USA |
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Abstract: | For the rapid establishment of optimal conditions for a genetic transformation system for tall fescue, several factors influencing transient gene expression were studied in protoplasts, after the reporter β-glucuronidase gene was introduced by electroporation. In a time-course study of transient gene expression, GUS activity peaked at 24 h after electroporation. Among the different field strength conditions tested, maximum GUS activity was observed at 750 V/cm. Increases in the amount of plasmid DNA to 80 μg/ml led to increased GUS activity. GUS activities increased in linear fashion with increasing protoplast densities up to 2 × 106/ml. Age of suspension cells from which protoplasts were derived influenced transient expression with maximum GUS activity obtained in 3- and 5-day-old suspensions. These results show that monocot and dicot protoplasts respond similarly in electroporation. |
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Keywords: | Festuca arundinacea Schreb Protoplasts Electroporation Transient expression assay |
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