Part:BBa_K1820015

CP11_RBS_sfGFP(Bs)_Terminator

Usage and Biology

This is a composite part designed as a report protein sequence for use in Lactococcus lactis. The promoter is a mid-range promoter from Peter Ruhdal Jensen and Karin Hammer's library of synthetic promoters for Lactococcus lactis followed by a popular ribosome binding site (Elowitz 1999), a super folded green fluorescent protein codon optimized for Bacillus subtilis (sfGFP(Bs)), and a popular double-stop terminator. Although it was designed for L. lactis, it has displayed function in Escherichia coli as well. The sfGFP(Bs) has worked well in a variety of bacteria, such as L. lactis, B. subtilis, Escherischia coli and Streptococcus pneumoniae and there are indications that the promoter is functional in a large number of prokaryotic organisms. As such, it is likely that this construct will be functional in a variety of prokaryotic organisms.

We created and tested this construct in Escherichia coli in the pSB1C3 plasmid. It was tested for fluorescence relative to non-transformed E. coli cell and two other similar constructs (see Figure 1).

Figure 1. Fluorescence levels from three constructs using synthetic promoters from Jensen and Hammer (1998) in E. coli excited at 485/20 with emissions read at 528/20. cp8 from part BBa_K1820014, cp11 from part BBa_K1820015, and cp44 from part BBa_K1820016

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Unknown

References

Jensen, P. R., Hammer, K. (1998). The Sequence of Spacers between the Consensus Sequences Modulates the Strength of Prokaryotic Promoters. Appl Environ Microbiol. 1998 Jan; 64(1): 82–87. http://www.ncbi.nlm.nih.gov/pmc/articles/PMC124675/

Overkamp, W. et al. (2013) Benchmarking various green fluorescent protein variants in Bacillus subtilis, Streptococcus pneumoniae, and Lactococcus lactis for live cell imaging. Appl. Environ. Microbiol. 79: 6481-6490