Difference between revisions of "Part:BBa K4164014"
 
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The composite part can be directly imported into plasmid and express FXR-ddRFPA1 induced with IPTG.
 
The composite part can be directly imported into plasmid and express FXR-ddRFPA1 induced with IPTG.
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We tried to determine the solubility of FXR-ddRFPA1 by purifying 6xHis-tagged proteins under native conditions and performing SDS-PAGE to compare the abundance of the band with the size of interest.Initially, FXR-ddRFPA1 is in the pellet after lysis and can't be purified(Lane1), then we optimized the expression conditions and purified it from supernatant of bacteria liquid.
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<p style="text-align: center;"><img src="https://static.igem.wiki/teams/4164/wiki/part-registry/sds-page.png"with="1000" height="" width="500" height=""/>
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<p style="text-align: center;"><b>Fig.1 Purification of FXR-ddRFPA1 and RXR-ddRFPB1.Lane 1: protein contained in the pellet after bacterial disruption. Lane 2: RXR-ddRFPB1 purified from supernatant of bacteria liquid. Lane 3: FXR-ddRFPA1 purified from supernatant of bacteria liquid in optimized expression conditon..</b></p>
  
 
<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 05:25, 12 October 2022


Inductive expression of FXR-ddRFPA1

In order to find an appropriate expression intensity to achieve balance between metabolic burden and detection efficiency, we also tried T7lac promoter from pET-29a+(Novagen).

The composite part can be directly imported into plasmid and express FXR-ddRFPA1 induced with IPTG.

We tried to determine the solubility of FXR-ddRFPA1 by purifying 6xHis-tagged proteins under native conditions and performing SDS-PAGE to compare the abundance of the band with the size of interest.Initially, FXR-ddRFPA1 is in the pellet after lysis and can't be purified(Lane1), then we optimized the expression conditions and purified it from supernatant of bacteria liquid.


Fig.1 Purification of FXR-ddRFPA1 and RXR-ddRFPB1.Lane 1: protein contained in the pellet after bacterial disruption. Lane 2: RXR-ddRFPB1 purified from supernatant of bacteria liquid. Lane 3: FXR-ddRFPA1 purified from supernatant of bacteria liquid in optimized expression conditon..

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 1558
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI.rc site found at 1147