Difference between revisions of "PGP1087"

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(The reference)
 
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'''General comments'''
 
'''General comments'''
  
Integrative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping the control of the expression under its natural promotor
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* Integrative [[plasmid]] for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping the control of the expression under its natural promotor, allows express of the downstream gene by the p(spac) promoter
* ''E.coli'': lowcopy plasmid, select for ampicillin
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* constructed in [[Jörg Stülke]]'s lab (C. Diethmaier)
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 +
* ''E.coli'': low copy plasmid, select for ampicillin
 
* ''B. subtilis'': no ori, select for erythromycin/lincomycin
 
* ''B. subtilis'': no ori, select for erythromycin/lincomycin
 +
 +
* based on pMUTIN-FLAG
  
 
'''Hints for the usage'''
 
'''Hints for the usage'''
 
* amplify the last 300bp of your gene of interest,  
 
* amplify the last 300bp of your gene of interest,  
* skip the Stop-Codon of the gene in the revers primer
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* skip the stop codon of the gene in the reverse primer
 
* take care that the integration is in-frame
 
* take care that the integration is in-frame
  
 
'''Antibody'''
 
'''Antibody'''
 
*dilute the anitbody 1:10000 (you can freeze the Blotto solution and reuse it)
 
*dilute the anitbody 1:10000 (you can freeze the Blotto solution and reuse it)
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=The reference=
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<pubmed>21856853</pubmed>
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==  Return to [[Plasmids]] ==

Latest revision as of 18:07, 2 August 2016

pGP1087: click to enlarge

General comments

  • Integrative plasmid for the fusion of 3x FLAG tag to the C-terminus of a protein, keeping the control of the expression under its natural promotor, allows express of the downstream gene by the p(spac) promoter
  • constructed in Jörg Stülke's lab (C. Diethmaier)
  • E.coli: low copy plasmid, select for ampicillin
  • B. subtilis: no ori, select for erythromycin/lincomycin
  • based on pMUTIN-FLAG

Hints for the usage

  • amplify the last 300bp of your gene of interest,
  • skip the stop codon of the gene in the reverse primer
  • take care that the integration is in-frame

Antibody

  • dilute the anitbody 1:10000 (you can freeze the Blotto solution and reuse it)

The reference

Christine Diethmaier, Nico Pietack, Katrin Gunka, Christoph Wrede, Martin Lehnik-Habrink, Christina Herzberg, Sebastian Hübner, Jörg Stülke
A novel factor controlling bistability in Bacillus subtilis: the YmdB protein affects flagellin expression and biofilm formation.
J Bacteriol: 2011, 193(21);5997-6007
[PubMed:21856853] [WorldCat.org] [DOI] (I p)

Return to Plasmids