Circadian Rhythms pp 115-129 | Cite as
Detection of Rhythmic Bioluminescence From Luciferase Reporters in Cyanobacteria
Abstract
The unicellular cyanobacterium Synechococcus elongatus PCC 7942 is the model organism for studying prokaryotic circadian rhythms. Although S. elongatus does not display an easily measurable overt circadian behavior, its gene expression is under circadian control; hence, a “behavior” is created by linking a cyanobacterial promoter to either the bacterial luxAB or firefly luc luciferase genes to create reporter fusions whose activity can be easily monitored by bioluminescence. Numerous vectors have been created in our lab for introducing luciferase reporter genes into the S. elongatus chromosome. A choice of methods and equipment to detect light production from the luciferase fusions provides a means for high-throughput, automated mutant screens as well as testing rhythms from two promoter fusions within the same cell culture.
Key Words
Synechococcus elongatus PCC 7942 cyanobacteria luciferase bioluminescence circadian rhythm neutral siteReferences
- 1.Golden, S. S., and Sherman, L. A. (1984) Optimal conditions for genetic transformation of the cyanobacterium Anacystis nidulans R2. J. Bacteriol. 158, 36–42.PubMedGoogle Scholar
- 2.Elhai, J., and Wolk, C. P. (1988) Conjugal transfer of DNA to cyanobacteria. Methods Enzymol. 167, 747–754.CrossRefPubMedGoogle Scholar
- 3.Golden, S. S., and Sherman, L. A. (1983) A hybrid plasmid is a stable cloning vector for the cyanobacterium Anacystis nidulans R2. J. Bacteriol. 155, 966–972.PubMedGoogle Scholar
- 4.Bustos, S. A., and Golden, S. S. (1991) Expression of the psbDII gene in Synechococcus sp. strain PCC 7942 requires sequences downstream of the transcription start site. J. Bacteriol. 173, 7525–7533.PubMedGoogle Scholar
- 5.Golden, S. S., Brusslan, J., and Haselkorn, R. (1987) Genetic engineering of the cyanobacterial chromosome. Methods Enzymol. 153, 215–231.CrossRefPubMedGoogle Scholar
- 6.Golden, S. S. (1988) Mutagenesis of cyanobacteria by classical and gene-transfer-based methods. Methods Enzymol. 167, 714–727.CrossRefPubMedGoogle Scholar
- 7.Sambrook, J., Fritsch, E. F., and Maniatis, T. (1989) Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, NY.Google Scholar
- 8.Liu, Y., Golden, S. S., Kondo, T., Ishiura, M., and Johnson, C. H. (1995) Bacterial luciferase as a reporter of circadian gene expression in cyanobacteria. J. Bacteriol. 177, 2080–2086.PubMedGoogle Scholar
- 9.Nair, U., Ditty, J. L., Min, H., and Golden, S. S. (2002) Roles for sigma factors in global circadian regulation of the cyanobacterial genome. J. Bacteriol. 184, 3530–3538.CrossRefPubMedGoogle Scholar
- 10.Golden, S. S., and Canales, S. R. (2003) Cyanobacterial circadian rhythms—timing is everything. Nature Rev. Microbiol. 1, 191–199.CrossRefGoogle Scholar
- 11.Plautz, J. D., Straume, M., Stanewsky, R., et al. (1997) Quantitative analysis of Drosophila period gene transcription in living animals. J. Biol. Rhythms. 12, 204–217.CrossRefPubMedGoogle Scholar
- 12.Aschoff, J. (1981) Freerunning and entrained circadian rhythms. In: Handbook of Behavioral Neurobiology: Biological Rhythms (Aschoff, J., ed.), Plenum Press, New York, pp. 81–93.Google Scholar
- 13.Ditty, J. L., Williams, S. B., and Golden, S. S. (2003) A cyanobacterial circadian timing mechanism. Annu. Rev. Genet. 37, 513–543.CrossRefPubMedGoogle Scholar
- 14.Kondo, T., and Ishiura, M. (1994) Circadian rhythms of cyanobacteria: monitoring the biological clocks of individual colonies by bioluminescence. J. Bacteriol. 176, 1881–1885.PubMedGoogle Scholar
- 15.Golden, S. S., and Haselkorn, R. (1985) Mutation to herbicide resistance maps within the psbA gene of Anacystis nidulans R2. Science 229, 1104–1107.CrossRefPubMedGoogle Scholar
- 16.Prentki, P., and Krisch, H. M. (1984) In vitro insertional mutagenesis with a selectable DNA fragment. Gene. 29, 303–313.CrossRefPubMedGoogle Scholar