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Applied and Environmental Microbiology, November 2004, p. 6809-6815, Vol. 70, No. 11
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.11.6809-6815.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.

Visualization of Differential Gene Expression by Improved Cyan Fluorescent Protein and Yellow Fluorescent Protein Production in Bacillus subtilis

Jan-Willem Veening, Wiep Klaas Smits, Leendert W. Hamoen,{dagger} Jan D. H. Jongbloed, and Oscar P. Kuipers*

Department of Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Haren, The Netherlands

Received 28 April 2004/ Accepted 24 June 2004

The distinguishable cyan and yellow fluorescent proteins (CFP and YFP) enable the simultaneous in vivo visualization of different promoter activities. Here, we report new cloning vectors for the construction of cfp and yfp fusions in Bacillus subtilis. By extending the N-terminal portions of previously described CFP and YFP variants, 20- to 70-fold-improved fluorescent-protein production was achieved. Probably, the addition of sequences encoding the first eight amino acids of the N-terminal part of ComGA of B. subtilis overcomes the slow translation initiation that is provoked by the eukaryotic codon bias present in the original cfp and yfp genes. Using these new vectors, we demonstrate that, within an isogenic population of sporulating B. subtilis cells, expression of the abrB and spoIIA genes is distinct in individual cells.

* Corresponding author. Mailing address: Department of Genetics, Groningen Biomolecular Sciences and Biotechnology Institute, University of Groningen, Kerklaan 30, 9751 NN Haren, The Netherlands. Phone: 31 50 3632093. Fax: 31 50 3632348. E-mail: o.p.kuipers{at}biol.rug.nl.

{dagger} Present address: Sir William Dunn School of Pathology, University of Oxford, Oxford OX1 3RE, United Kingdom.

Applied and Environmental Microbiology, November 2004, p. 6809-6815, Vol. 70, No. 11
0099-2240/04/$08.00+0     DOI: 10.1128/AEM.70.11.6809-6815.2004
Copyright © 2004, American Society for Microbiology. All Rights Reserved.



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