Expression and Secretion of Cyan Fluorescent Protein (CFP) in B. subtilis using the Chitinase Promoter from Bacillus pumilus SG2

Authors
Abbas Shali 1
Garshasb Rigi 2
Majid Pornour 3
Gholamreza Ahmadian 1
1 Department of Industrial and Environmental Biotechnology, National Institute of Genetic Engineering and Biotechnology (NIGEB) , P.O. Box 14155-6343 , Tehran, Iran
2 Department of Biology, Faculty of Science, Behbahan Khatam Alanbia University of Technology, Behbahan, Khuzestan, Iran
3 Medical Laser Research Center, Academic Center for Education, Culture and Research (ACECR), Tehran, Iran
10.18869/acadpub.ibj.21.4.240
Abstract
Background: Improved cyan fluorescent protein (ICFP) is a monochromic, green fluorescent protein (GFP) derivative produced by Aequorea macrodactyla in a process similar to GFP. This protein has strong absorption spectra at wavelengths 426-446 nm. ICFP can be used in cell, organelle or intracellular protein labeling, investigating the protein-protein interactions as well as assessing the promoter activities. Methods: In our previous study, the promoters of two chitinases (ChiS and ChiL) from Bacillus pumilus SG2 were assessed in B. subtilis and their regulatory elements were characterized. In the present study, icfp was cloned downstream of several truncated promoters obtained in the former study, and ICFP expression was evaluated in B. subtilis. Results: Extracellular expression and secretion of ICFP were analyzed under the control of different truncated versions of ChiSL promoters grown on different media. Results from SDS-PAGE and fluorimetric analyses showed that there were different expression rates of CFP; however, the UPChi-ICFP3 construct exhibited a higher level of expression and secretion in the culture medium. Conclusion: Our presented results revealed that inserting this truncated form of Chi promoter upstream of the ICFP, as a reporter gene, in B. subtilis led to an approximately ten fold increase in ICFP expression.
Keywords
Cyan fluorescent protein
Medical reporter gene
Medical screenable marker
Medicinal Protein Engineering