Difference between revisions of "Part:BBa K847210:Design"
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Potential cell-cycle dependent promoters in ''Escherichia coli'' and their sequences were identified from literature (Quiñones 1997) and isolated using Colony PCR. We developed our constructs for assay by ligating these PCR products in front of [[Part:BBa_E0840|BBa_E0840]], which consists of RBS [[Part:BBa_B0030|BBa_B0030]], reporter [[Part:BBa_E0040|BBa_E0040]] (mut3b GFP), and terminator [[Part:BBa_B0015|BBa_B0015]]. | Potential cell-cycle dependent promoters in ''Escherichia coli'' and their sequences were identified from literature (Quiñones 1997) and isolated using Colony PCR. We developed our constructs for assay by ligating these PCR products in front of [[Part:BBa_E0840|BBa_E0840]], which consists of RBS [[Part:BBa_B0030|BBa_B0030]], reporter [[Part:BBa_E0040|BBa_E0040]] (mut3b GFP), and terminator [[Part:BBa_B0015|BBa_B0015]]. | ||
| − | This construct functions in ''E. coli'' | + | This construct functions in ''E. coli'' but may also work in prokaryotes with similar DnaA dynamics (e.g. Caulobacter). |
===Source=== | ===Source=== | ||
Latest revision as of 22:23, 3 October 2012
polA (DNA Polymerase I) DnaA-activated promoter
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 144
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Design Notes
Potential cell-cycle dependent promoters in Escherichia coli and their sequences were identified from literature (Quiñones 1997) and isolated using Colony PCR. We developed our constructs for assay by ligating these PCR products in front of BBa_E0840, which consists of RBS BBa_B0030, reporter BBa_E0040 (mut3b GFP), and terminator BBa_B0015.
This construct functions in E. coli but may also work in prokaryotes with similar DnaA dynamics (e.g. Caulobacter).
Source
E. coli strain K12
References
Messer M., Weigel C.: DnaA initiator - also a transcription factor. Molecular Microbiology 1997, 24:1-6.
Chiaramello A.E., Zyskind, J.W.: Coupling of DNA replication to growth rate in Escherichia coli: A possible role for guanosine tetraphosphate. J. Bacteriology 1990, 172:2013-2019.
Quiñones A., Wandt G., Kleinstauber S., Messer W.: DnaA protein stimulates polA gene expression in Escherichia coli. Molecular Microbiology 1997, 23: 1193-1202.
Messer W.: The bacterial replication initiator DnaA. DnaA and oriC, the bacterial mode to initiate DNA replication 2002, 26:355-374.
Lam A., St-Pierre F., Gong Y., Marshall J.D., Cranfill P.J., Baird M.A., McKeown M.R., Wiedenmann J., Davidson M.W., Schnitzer M., Tsien R.Y., Lin M.Z.: Improved dynamic range of genetically encoded FRET sensors with bright new green and red fluorescent proteins. Nature Methods 2012, 9:1005-1012.
Ferullo D.J., Cooper D.L., Moore H.R., Lovett S.T.: Cell cycle synchronization of E. coli using the stringent response, with fluorescence labeling assays for DNA content and replication. Methods 2009, 48:8-13.
Grant M.A.A., Saggioro C., Ferrari U., Bassetti B., Sclavi B., Lagomarsino M.C.: DnaA and the timing of chromosome replication in Escherichia coli as a function of growth rate. BMC Systems Biology 2011, 5:201.