Difference between revisions of "Part:BBa K1412014"

Revision as of 09:03, 6 October 2014

Characterize the efficiency of pTetR (R0040) with chemotaxis

BBa_K1412014: pTet-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. Then we can characterize the efficiency of promoter via measuring the migration distance positively associated with the expression strength of CheZ.

Usage

When we want to characterize the efficiency of promoter, we usually link the promoter with GFP, then characterize the promoter by just measure the fluorescence intensity of GFP. In our part, you need just link RBS(1.0) after a pTetR promoter and before a CheZ gene, ending with a TT terminator(pTetR-RBS(changeable)-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

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
COMPATIBLE 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.

3.Next, link RBS-CheZ-TT with promoter pTetR.

Characterization stage

1.Transfer the part pTetR-RBS-CheZ-TT into E.coli (CheZ knock out), and coat plates, culture for hours to measure the migration diameter of E.coli.

@@ Line 15: / Line 15: @@
 ----
-When we want to characterize the efficiency of RBS, we usually link the RBS with GFP, then characterize the RBS by just measure the fluorescence intensity of GFP. In our part, you need just link RBS after a Plac promoter and before a <i>CheZ</i> gene, ending with a TT terminator(Plac-RBS(changeable)-<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>.
+When we want to characterize the efficiency of promoter, we usually link the promoter with GFP, then characterize the promoter by just measure the fluorescence intensity of GFP. In our part, you need just link RBS(1.0) after a pTetR promoter and before a <i>CheZ</i> gene, ending with a TT terminator(pTetR-RBS(changeable)-<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>.
@@ Line 69: / Line 69: @@
 .Then the skeleton <i>CheZ</i>-TT link to RBS.
-.Next, link  RBS-<i>CheZ</i>-TT with promoter Plac.
+.Next, link  RBS-<i>CheZ</i>-TT with promoter pTetR.
 ===='''Characterization stage'''====
-.Transfer the part Plac-RBS-<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>.
+.Transfer the part pTetR-RBS-<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>.