Promoter

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Protoplast Isolation And Transient Gene Expression In Switchgrass, Panicum Virgatum L.

Journal Title, Volume, Page: 
Biotechnology Journal 2008, 3, 354–359
Year of Publication: 
2008
Authors: 
Mitra Mazarei
Department of Plant Sciences, University of Tennessee, Knoxville, TN, USA
Hani Al-Ahmad
Department of Plant Sciences, University of Tennessee, Knoxville, TN, USA
Current Affiliation: 
Department of Biology & Biotechnology, Faculty of Science, An-Najah National University, Nablus. Palestine
Mary R. Rudis
Department of Plant Sciences, University of Tennessee, Knoxville, TN, USA
C. Neal Stewart, Jr
Department of Plant Sciences, University of Tennessee, Knoxville, TN, USA
Preferred Abstract (Original): 

Transient assay systems using protoplasts have been utilized in several plant species and are a powerful tool for rapid functional gene analysis and biochemical manipulations. A protoplast system has not been used in switchgrass (Panicum virgatum L.), even though it is a bioenergy crop that has received considerable attention. Here we report the first protocol to isolate large numbers of viable protoplasts from both leaves and roots of two switchgrass genotypes. Furthermore, we demonstrate transient expression of PEG-mediated DNA uptake in the isolated protoplasts by measuring the activity of β-glucuronidase (GUS) reporter gene driven by either the Cauliflower mosaic virus (CaMV) 35S promoter or the maize ubiquitin 1 promoter. Protoplast transformation with either the 35S or the ubiquitin promoter resulted in an increase in GUS activity compared to the untransformed controls; however, the extent of GUS activity was considerably higher for the ubiquitin promoter than for the 35S promoter. Collectively, our results indicate an efficient protoplast isolation and transient assay system that can be used to facilitate gene expression studies in switchgrass.

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