Product CartCyan-Green Fluorescent Proteins (for anaerobic and aerobic environments)
• Bright cyan-green fluorescence
• No need for oxygen as cofactor
• Can be used in anaerobic and semi-anaerobic environments
Fluorescent reporter proteins are valuable noninvasive molecular tools for in vivo real-time imaging of living cells and tissues as well as in vitro fluorescence labeling.
The flavin mononucleotide (FMN)-based fluorescent proteins (FbFPs) from evocatal´s evoglow series were developed to work in anaerobic biological systems. In contrast to GFP-like reporter proteins such as YFP (see Fig. 1) the evoglow reporter proteins do not need oxygen as a cofactor for the synthesis of their respective chromophores. This enables the application in anaerobic and semi-anaerobic environments.
Now you can finally use in vivo fluorescent reporter technology to explore microbial biofilms and other cells growing under anaerobic or semiaerobic conditions.
Microbial biofilms are the most common mode of bacterial growth in nature and are highly resistant to antibiotics (relevant in clinical infections). This form of microbial growth is being studied in a wide range of scientific disciplines including biomedicine, water engineering and evolutionary biology.
In vivo-fluorescence of evoglow-protein PpFbFP compared to that of conventional Yellow Fluorescent Protein (YFP).
Comparison of YFP and PpFbFP-mediated in vivo fluorescence in the absence and presence of oxygen was carried out in the facultative anaerobic bacterium Rhodobacter capsulatus. R. capsulatus B10S-T7 cells carrying an expression plasmid encoding either YFP or Pp1 were grown under aerobic as well as anaerobic conditions and in vivo fluorescence was analyzed microscopically using a Zeiss LSM 510 confocal laser scanning microscope. Cells were excited with laser light at defined wavelengths of 488 nm (YFP) and 458 nm (Pp1). Fluorescence emission was detected in the ranges of 505–550 nm (YFP) and 475–525 nm (Pp1), respectively.
Control experiments with R. capsulatus cells without a fluorescent reporter gene were performed in the same way.
Three different Flavin Mononucleotide (FMN)-based fluorescent proteins, Bs1, Bs2, and Pp1, all with an excitation maximum at 450 nm, an emmission maximum of 495 nm, and all showing no toxicity, are available:
Bs1
Bs1 is derived from the Bacillus subtilis blue-light photoreceptor YtvA and has the B. subtilis wildtype sequence.
The molecular weight is only 33 kDa and the protein is a monomer in solution.
Bs2
Bs2 carries the DNA sequence of a truncated YtvA gene encoding only the photoactive LOV domain and optimized for codon usage in E. coli. The molecular weight is just 19 kDA. The protein forms homodimers in solution.
Pp1
Pp1 is derived from the Pseudomonas putida gene SB2. The molecular weight is also 19 kDA, and the protein forms homodimers in solution as well.
All three proteins are available as cDNA harbouring plasmids (individually or as a set) or expression ready broad gram negative host range vectors with inducible or constitutive FP driving promoter (individually or as sets).
E. coli cells harbouring Pp1 expression plasmid.
Left: Halogen Incident Light, Right: Fluorescence Microscopy
Reference
Drepper, T. et al. (2007) Reporter proteins for in vivo fluorescence without oxygen. Nature Biotechnology 25: 443-445.
Nature Biotechnology Publication
User Manual for cDNA Vectors
User Manual for Expression Vectors
