Difference between revisions of "Part:BBa K4214003"

 
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<partinfo>BBa_K4214003 short</partinfo>
 
<partinfo>BBa_K4214003 short</partinfo>
  
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LacI (BBa_J04500) and Human GIF with His6-tag. This part is a composite part from BBa_K4214002 with the addition of LacI sequence(BBa_J04500). The LacI sequence is used in order to induce our protein expression with IPTG.  
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LacI [https://parts.igem.org/Part:BBa_J04500 (BBa_J04500)] and Human GIF with His6-tag. This part is a composite part from BBa_K4214002 with the addition of LacI sequence [https://parts.igem.org/Part:BBa_J04500 (BBa_J04500)]. The LacI sequence is used in order to induce our protein expression with IPTG, and the 6xHis-tag was used for protein purification.
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[[Image:BBa K4214003 Lac-IF-His.png|250px|thumb|right|Fig.: ''LacI (BBa_J04500) and Human GIF with His6-tag (BBa_K4214002) '']]
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=Usage and Biology=
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<hr>
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*NCBI page of the human gastric intrinsic factor [https://www.ncbi.nlm.nih.gov/gene/2694]
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*This part of hGIF does not contain the native signal peptide (AA1-18)
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*The LacI promotor is described in part [https://parts.igem.org/Part:BBa_J04500 BBa_J04500]
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=Characterization=
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<hr>
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===SDS-PAGE===
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A production culture was set up using BL21 ''E. coli'' and induced with 0.5 mM IPTG once their ODs reached 0.5. The cultures were left overnight at 30°C for protein production. The cells were then harvested, pelleted and purified using a HisPur Cobalt resin. The SDS-PAGE analysis of the production process is shown on the right.
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===Usage and Biology===
 
  
 
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Revision as of 13:19, 9 October 2022


hGIF with LacI promotor and 6xHis-tag

LacI (BBa_J04500) and Human GIF with His6-tag. This part is a composite part from BBa_K4214002 with the addition of LacI sequence (BBa_J04500). The LacI sequence is used in order to induce our protein expression with IPTG, and the 6xHis-tag was used for protein purification.

Fig.: LacI (BBa_J04500) and Human GIF with His6-tag (BBa_K4214002)


Usage and Biology

  • NCBI page of the human gastric intrinsic factor [1]
  • This part of hGIF does not contain the native signal peptide (AA1-18)
  • The LacI promotor is described in part BBa_J04500

Characterization

SDS-PAGE

A production culture was set up using BL21 E. coli and induced with 0.5 mM IPTG once their ODs reached 0.5. The cultures were left overnight at 30°C for protein production. The cells were then harvested, pelleted and purified using a HisPur Cobalt resin. The SDS-PAGE analysis of the production process is shown on the right.



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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 485
    Illegal BsaI.rc site found at 1165
    Illegal SapI site found at 1155