BIOL 192 |
| Gene reporter systems have been an important tool to study the regulation of gene expression. The most common gene reporter has traditionally been the lacZ gene from E.coli. Unfortunately, this reporter cannot be used in Candida albicans because the codon CUG is read by tRNA in C.albicans as serine instead of leucine, resulting in a nonfunctional enzyme. The Renilla reniformis luciferase gene does not contain CUG codons and produces an enzyme that catalyzes the conversion of its substrate coelenterazine to coelenteramide, releasing light that can be quantified using a luminometer. We present the construction of six different reporter plasmids containing three selectable markers (URA3, HIS1, and ARG4) for C. albicans, and a large restriction site polylinker in two different orientations. We demonstrate the optimum conditions for cell lysis, assaying activity, assay reliability, and the in vivo half life of the luciferase protein in C. albicans. |
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Frontiers in Chemical Biology
5:00 PM-7:00 PM, Wednesday, August 22, 2007 BCEC -- Exhibit Hall - B2, Poster
Division of Biological Chemistry |