Difference between revisions of "Part:BBa K3114014"

 
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<partinfo>BBa_K3114014 short</partinfo>
 
<partinfo>BBa_K3114014 short</partinfo>
  
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===Usage and Biology===
 
===Usage and Biology===
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This part can be used for fusion to the C-terminal of proteins including the ICARUS spacer and a double terminator. ICARUS can be used for the purification of large proteins that have strong electrostatic forces within their cores, using a 6xHis tag (Ni-NTA column purification). ICARUS contains flanking SacII restriction sites for removal at the DNA level and a thrombin proteolytic site for protein cleavage.
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This part consists of the double terminator BBa_B0014 that has been modified to be compatible with the MoClo standard for Golden Gate assembly (Weber et al., 2011). 
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[[Image:T--Calgary--Repurposing-Chlorophyll-Constructs-Design.png|500px|thumb|right|Figure 1. iGEM Calgary's genetic construct scheme for chlorophyll degradation pathway enzymes. Each construct consists of a T7 inducible promoter, a strong RBS, one of four chlorophyll degradation pathway enzymes, a novel spacer containing a 6xHis tag (ICARUS), and a bidirectional terminator.]]
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iGEM Calgary engineered its chlorophyll degradation pathway enzymes to contain this ICARUS spacer to allow for successful protein purification. These parts can be found in our collection [https://2019.igem.org/Team:Calgary/Part_Collection collection.]
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<ul>
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<li>Chlorophyll B Reductase (CBR) inducible circuit [https://parts.igem.org/Part:BBa_K3114024 (BBa_K3114024)]</li>
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<li>7-Hydroxymethyl Chlorophyll A Reductase (7-HCAR) inducible circuit [https://parts.igem.org/Part:BBa_K3114025 (BBa_K3114025)]</li>
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<li>Magnesium Dechelatase Stay Green (SGR) inducible circuit [https://parts.igem.org/Part:BBa_K3114026 (BBa_K3114026)]</li>
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<li>Pheophytinase (PPH) inducible circuit [https://parts.igem.org/Part:BBa_K3114027 (BBa_K3114027)]</li>
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</ul>
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The ICARUS sequence was designed as a novel contribution to the iGEM community. ICARUS itself can be tested for its use in fusion proteins or for the purification of other proteins that have previously not been purified using a 6xHis tag.
  
 
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Revision as of 20:29, 12 December 2019


ICARUS spacer + 6XHis tag + double terminator for purification of highly electronegative proteins

Usage and Biology

This part can be used for fusion to the C-terminal of proteins including the ICARUS spacer and a double terminator. ICARUS can be used for the purification of large proteins that have strong electrostatic forces within their cores, using a 6xHis tag (Ni-NTA column purification). ICARUS contains flanking SacII restriction sites for removal at the DNA level and a thrombin proteolytic site for protein cleavage.

This part consists of the double terminator BBa_B0014 that has been modified to be compatible with the MoClo standard for Golden Gate assembly (Weber et al., 2011).

File:T--Calgary--Repurposing-Chlorophyll-Constructs-Design.png
Figure 1. iGEM Calgary's genetic construct scheme for chlorophyll degradation pathway enzymes. Each construct consists of a T7 inducible promoter, a strong RBS, one of four chlorophyll degradation pathway enzymes, a novel spacer containing a 6xHis tag (ICARUS), and a bidirectional terminator.

iGEM Calgary engineered its chlorophyll degradation pathway enzymes to contain this ICARUS spacer to allow for successful protein purification. These parts can be found in our collection collection.

The ICARUS sequence was designed as a novel contribution to the iGEM community. ICARUS itself can be tested for its use in fusion proteins or for the purification of other proteins that have previously not been purified using a 6xHis tag.

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 215
    Illegal XhoI site found at 318
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 1
    Illegal BsaI.rc site found at 321