Difference between revisions of "Part:BBa K1949000"
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This new promoter, a cold inducible promoter (we call this Pcold) consists of the cspA promoter, Cold Box, 5’-UTR, RBS and DB. This promoter is used to effectively produce proteins at low temperatures. | This new promoter, a cold inducible promoter (we call this Pcold) consists of the cspA promoter, Cold Box, 5’-UTR, RBS and DB. This promoter is used to effectively produce proteins at low temperatures. | ||
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<i>E. coli</i> cells which carry the P<i>tet</i>-RBS-<i>gfp</i> plasmid was cultured at 18℃, and florescence intensity was measured at indicated time points. Also, the same experiment was performed at 37℃. We thought this was because <i>gfp</i> is easy to fold for florescent at low temperature. By contrast, florescence of <i>E. coli</i> which included P<i>tet</i>-RBS-<i>gfp</i> at 18℃ was about eight times as florescence as it at 37℃. By this result, we confirm P<i>cold</i> activates gene expression at low temperature. | <i>E. coli</i> cells which carry the P<i>tet</i>-RBS-<i>gfp</i> plasmid was cultured at 18℃, and florescence intensity was measured at indicated time points. Also, the same experiment was performed at 37℃. We thought this was because <i>gfp</i> is easy to fold for florescent at low temperature. By contrast, florescence of <i>E. coli</i> which included P<i>tet</i>-RBS-<i>gfp</i> at 18℃ was about eight times as florescence as it at 37℃. By this result, we confirm P<i>cold</i> activates gene expression at low temperature. | ||
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===Biobrick Tips=== | ===Biobrick Tips=== | ||
Revision as of 10:35, 10 October 2016
cold inducible promoter (Pcold)
This new promoter, a cold inducible promoter (we call this Pcold) consists of the cspA promoter, Cold Box, 5’-UTR, RBS and DB. This promoter is used to effectively produce proteins at low temperatures.
Characterization
We measured florescence intensity per turbidity using cells cultured at 18℃ and 37℃ to confirm function of Pcold. The cells had a plasmid which carries Pcold-gfp or Ptet-gfp.
E. coli cells which carry the Ptet-RBS-gfp plasmid was cultured at 18℃, and florescence intensity was measured at indicated time points. Also, the same experiment was performed at 37℃. We thought this was because gfp is easy to fold for florescent at low temperature. By contrast, florescence of E. coli which included Ptet-RBS-gfp at 18℃ was about eight times as florescence as it at 37℃. By this result, we confirm Pcold activates gene expression at low temperature.
Biobrick Tips
This part is not able to be used for most common assembly, because restriction enzyme digestion with XbaI and SpeI generates an unexpected stop codon. Therefore, this part do not meet the criteria of basic parts construction. Our team generated a unique digestion site, BamHI at the upstream of the suffix. We recommend to use this BamHI site for cloning.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 308
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]