Difference between revisions of "Part:BBa K1412801"
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3.Next, connect  RBS-<i>CheZ</i>-TT with promoter pLac.
 
3.Next, connect  RBS-<i>CheZ</i>-TT with promoter pLac.
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===='''Characterization stage'''====
 
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1.Transfer the part pLac-RBS-<i>CheZ</i>-TT into <i>E. coli</i> (<i>CheZ</i> knocked out), and coat plates, culture for hours to measure the migration diameter of <i>E. coli</i>.
 
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[[File:Example.jpg |500px]]
 
  
 
==='''Reference'''===
 
==='''Reference'''===

Revision as of 21:14, 17 October 2014

Characterize efficiency of RBS with chemotaxis

BBa_K1412801: pLac-RBS(0.01)-CheZ-TT

This part consists of [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 RBS or promoter efficiency by replacing different promoters or ribosome binding sites(RBS).Then we can characterize the efficiency of RBS and promoter via measuring the migration distance positively associated with the expression strength of CheZ.

Characterization.jpg


Usage

When we want to characterize the efficiency of RBS, we usually connect the RBS with GFP, then measuring the fluorescence intensity of GFP. In our part, you need just connect RBS after a pLac promoter and before a CheZ gene, ending with double terminator(pLac-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

BBa_K1412000: pLac-RBS(1.0)-CheZ-TT CheZ generator under pLac promoter

BBa_K1412005: RBS(1.0)-CheZ-TT

BBa_K1412006: RBS(0.01)-CheZ-TT

BBa_K1412007: RBS(0.3)-CheZ-TT

BBa_K1412014: pTet-RBS(1.0)-CheZ-TT Characterize the efficiency of promoters with chemotaxis

BBa_K1412614: pBAD-RBS(1.0)-CheZ-TT Characterize the efficiency of promoters with chemotaxis

BBa_K1412829: Plac-RBS(0.3)-CheZ-TT Characterize efficiency of RBS with chemotaxis

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 connect stage

1.As a backbone, double terminator connect with target gene CheZ.

2.Then connect the backbone RBS(0.01)to target gene CheZ-TT.

3.Next, connect RBS-CheZ-TT with promoter pLac.

Reference

More information, click here: [http://2014.igem.org/Team:XMU-China# XMU-China]