Difference between revisions of "Part:BBa K1412614"
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---- | ---- | ||
− | When we want to characterize the efficiency of | + | When we want to characterize the efficiency of promoter, we usually link the promoter with GFP, then characterize the efficiency of promoter by just measure the fluorescence intensity of GFP. In our part, you need just link RBS(1.0) after a pBAD promoter and before a <i>CheZ</i> gene, ending with a TT terminator(pBAD-RBS(1.0)-<i>CheZ</i>-TT). Then transfer this gene circuit into <i>E.coli</i> (<i>CheZ</i> knocked out), and coat plates, culuture on semi-solid medium to measure the migration diameter of <i>E.coli</i>. |
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1.As a skeleton, TT terminator link with <i>CheZ</i>. | 1.As a skeleton, TT terminator link with <i>CheZ</i>. | ||
− | 2.Then the skeleton <i>CheZ</i>-TT link to RBS. | + | 2.Then the skeleton <i>CheZ</i>-TT link to RBS(1.0). |
− | 3.Next, link RBS-<i>CheZ</i>-TT with promoter | + | 3.Next, link RBS-<i>CheZ</i>-TT with promoter pBAD. |
===='''Characterization stage'''==== | ===='''Characterization stage'''==== | ||
− | 1.Transfer the part | + | 1.Transfer the part pBAD-RBS(1.0)-<i>CheZ</i>-TT into <i>E.coli</i> (<i>CheZ</i> knock out), and coat plates, culture for hours to measure the migration diameter of <i>E.coli</i>. |
Revision as of 09:08, 6 October 2014
Characterize the efficiency of promoter(
BBa_K1412614: pBAD-RBS(1)-CheZ-TT
This part consists of a [http://en.wikipedia.org/wiki/Chemotaxis CheZ] gene which can express CheZ protein deciding E.coli whether tumble or swim straight. In this light, we can characterize the efficiency of promoter by just change different promoters before a CheZ gene. Then we can characterize the efficiency of promoter via measuring the migration distance positively associated with the expression strength of CheZ protein.
Usage
When we want to characterize the efficiency of promoter, we usually link the promoter with GFP, then characterize the efficiency of promoter by just measure the fluorescence intensity of GFP. In our part, you need just link RBS(1.0) after a pBAD promoter and before a CheZ gene, ending with a TT terminator(pBAD-RBS(1.0)-CheZ-TT). Then transfer this gene circuit into E.coli (CheZ knocked out), and coat plates, culuture on semi-solid medium to measure the migration diameter of E.coli.
Relevant parts
Notes
Source
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 125
- 21INCOMPATIBLE WITH RFC[21]Illegal BamHI site found at 65
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Protocol
Genetic link stage
1.As a skeleton, TT terminator link with CheZ.
2.Then the skeleton CheZ-TT link to RBS(1.0).
3.Next, link RBS-CheZ-TT with promoter pBAD.
Characterization stage
1.Transfer the part pBAD-RBS(1.0)-CheZ-TT into E.coli (CheZ knock out), and coat plates, culture for hours to measure the migration diameter of E.coli.
Reference