Difference between revisions of "Part:BBa K1820014"
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<p> 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.</p> | <p> 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.</p> | ||
− | <p>We created and tested this construct in ''Escherichia coli'' in the pSB1C3 plasmid. It was tested for fluorescence relative to non-transformed ''E. coli'' | + | <p>We created and tested this construct in ''Escherichia coli'' in the pSB1C3 plasmid. It was tested for fluorescence relative to non-transformed ''E. coli'' cells (see Figure 1).</p> |
Revision as of 23:32, 18 September 2015
CP8_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 cells (see Figure 1).
Figure 1. Fluorescence levels from this construct in E. coli excited at 485/20 with emissions read at 528/20
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000INCOMPATIBLE WITH RFC[1000]Illegal SapI.rc site found at 99
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