| Title: | A family of destabilized cyan fluorescent proteins as transcriptional reporters in S. cerevisiae |
| Author(s): | Hackett EA; Esch RK; Maleri S; Errede B; |
| Address: | "Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, NC 27599-7260, USA" |
| ISSN/ISBN: | 0749-503X (Print) 0749-503X (Linking) |
| Abstract: | "The 'programmable' features of the N-end rule degradation pathway and a ubiquitin fusion strategy were exploited to create a family of destabilized cyan fluorescent proteins (CFP) to be used as transcriptional reporters. The N-degron CFP reporters characterized in this report have half-lives of approximately 75, 50 and 5 min, but further modification of the N-degron signal sequences could readily generate additional variants within this range. These destabilized CFP reporters have been engineered into convenient plasmid constructs with features to enable their expression from upstream activating sequences of choice and to facilitate their targeted integration to the URA3-TIM9 intergenic region of chromosome V. The advantages and limitations of these reporters as temporal indicators of gene expression in living cells are illustrated by their application as reporters of galactose- and pheromone-induced transcription. The plasmid design we describe and the range of different stabilities that are theoretically feasible with this strategy make the N-degron CFP reporters easily adapted to a variety of applications" |
| Keywords: | "Amino Acid Motifs Amino Acid Sequence Blotting, Northern Blotting, Western Chromosomes, Fungal/genetics DNA, Fungal/genetics Fluorescent Dyes/chemistry/metabolism Galactokinase/genetics Gene Expression Regulation, Fungal Genes, Reporter/*genetics/physiolo;" |
| Notes: | "MedlineHackett, Elizabeth A Esch, R Keith Maleri, Seth Errede, Beverly eng R01 GM067809/GM/NIGMS NIH HHS/ GM067809/GM/NIGMS NIH HHS/ GM39852/GM/NIGMS NIH HHS/ Research Support, N.I.H., Extramural England 2006/04/07 Yeast. 2006 Apr 15; 23(5):333-49. doi: 10.1002/yea.1358" |
