Difference between revisions of "Part:BBa K525123"
(→‎Expression in E. coli)
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===Expression in ''E. coli''===
 
===Expression in ''E. coli''===
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The CspB gen was fused with a monomeric RFP ([https://parts.igem.org/Part:BBa_E1010 BBa_E1010]) using [http://2011.igem.org/Team:Bielefeld-Germany/Protocols#Gibson_assembly Gibson assembly] for characterization.
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For characterization the CspB gen was fused with a monomeric RFP ([https://parts.igem.org/Part:BBa_E1010 BBa_E1010]) using Gibson assembly.
  
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The CspB|mRFP fusion protein was overexpressed in E. coli KRX after induction of T7 polymerase by supplementation of 0,1 % L-rhamnose using the [http://2011.igem.org/Team:Bielefeld-Germany/Protocols/Downstream-processing#Expression_of_S-layer_genes_in_E._coli autinduction protocol] from promega.
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The CspB|mRFP fusion protein was overexpressed in E. coli KRX after induction of T7 polymerase by supplementation of 0,1 % L-rhamnose using the autinduction protocol from promega.
  
 
[[Image:Bielefeld_2011_BF3_Growthcurve.png|600px|center|thumb| '''Figure 1: Growthcurve of ''E. coli'' KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.''']]
 
[[Image:Bielefeld_2011_BF3_Growthcurve.png|600px|center|thumb| '''Figure 1: Growthcurve of ''E. coli'' KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.''']]
  
 
[[Image:Bielefeld_2011_BF3_RFU_OD.png|600px|center|thumb| '''Figure 2: RFU to OD<sub>600</sub> ratio of ''E. coli'' KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.''']]
 
[[Image:Bielefeld_2011_BF3_RFU_OD.png|600px|center|thumb| '''Figure 2: RFU to OD<sub>600</sub> ratio of ''E. coli'' KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.''']]

Revision as of 18:53, 21 September 2011

S-layer cspB from Corynebacterium glutamicum with lipid anchor and PT7 and RBS


Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 1334
    Illegal XhoI site found at 161
    Illegal XhoI site found at 779
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 1313
    Illegal SapI site found at 560
    Illegal SapI site found at 772
    Illegal SapI site found at 1320


Expression in E. coli

For characterization the CspB gen was fused with a monomeric RFP (BBa_E1010) using Gibson assembly.

The CspB|mRFP fusion protein was overexpressed in E. coli KRX after induction of T7 polymerase by supplementation of 0,1 % L-rhamnose using the autinduction protocol from promega.

Figure 1: Growthcurve of E. coli KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.
Figure 2: RFU to OD600 ratio of E. coli KRX expressing the fusion protein of CspB and mRFP with and without induction. A curve depicting KRX wildtype is shown for comparsion.