Difference between revisions of "Part:BBa K4047039"

 
 
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<partinfo>BBa_K4047039 short</partinfo>
 
<partinfo>BBa_K4047039 short</partinfo>
  
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Plasmid for deletion of fructose-1,6-bisphosphatase/ sedoheptulose-1,7-bisphosphatase in Synechococcus elongatus PCC 7942; contains all the components of pXWK3-glgC in addition to our gene
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This plasmid is for the deletion of glpX, encoding a bifunctional enzyme for fructose-1,6-bisphosphatase (FBPase) and sedoheptulose-1,7-bisphosphatase (SBPase) in Synechococcus elongatus PCC 7942. This plasmid was successfully integrated into transforming cells of this strain, detailed on the experience page, but was not able to achieve complete segregation (transformation of all genome copies) due to the essential nature of SBPase functionality for the cell. Successful integration was verified with PCR.
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This plasmid inserts kanamycin resistance (<partinfo>BBa_K4047019</partinfo>) into the target gene, preventing functional enzyme synthesis and providing kanamycin resistance as a selective marker. This plasmid is built from the plasmid backbone <partinfo>BBa_K4047018</partinfo>, which also contains gentamicin resistance (<partinfo>BBa_K4047016</partinfo>).
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Although this part lacks standard compatibility due to the enzymes available to our team, the plasmid does successfully integrate into the S. elongatus genome for the deletion of SBPase. We strongly recommend that future iGEM teams optimize this plasmid for compatibility and future work in engineering of the Calvin cycle.
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<!-- Add more about the biology of this part here
 
<!-- Add more about the biology of this part here

Latest revision as of 18:04, 20 October 2021


pGEM-SBPase (plasmid for deletion of SBPase)

This plasmid is for the deletion of glpX, encoding a bifunctional enzyme for fructose-1,6-bisphosphatase (FBPase) and sedoheptulose-1,7-bisphosphatase (SBPase) in Synechococcus elongatus PCC 7942. This plasmid was successfully integrated into transforming cells of this strain, detailed on the experience page, but was not able to achieve complete segregation (transformation of all genome copies) due to the essential nature of SBPase functionality for the cell. Successful integration was verified with PCR.

This plasmid inserts kanamycin resistance (BBa_K4047019) into the target gene, preventing functional enzyme synthesis and providing kanamycin resistance as a selective marker. This plasmid is built from the plasmid backbone BBa_K4047018, which also contains gentamicin resistance (BBa_K4047016).

Although this part lacks standard compatibility due to the enzymes available to our team, the plasmid does successfully integrate into the S. elongatus genome for the deletion of SBPase. We strongly recommend that future iGEM teams optimize this plasmid for compatibility and future work in engineering of the Calvin cycle.


Sequence and Features


Assembly Compatibility:
  • 10
    INCOMPATIBLE WITH RFC[10]
    Illegal PstI site found at 1998
    Illegal PstI site found at 2591
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal PstI site found at 1998
    Illegal PstI site found at 2591
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 1489
    Illegal BglII site found at 4478
    Illegal BamHI site found at 2345
    Illegal XhoI site found at 2433
    Illegal XhoI site found at 3675
    Illegal XhoI site found at 4474
  • 23
    INCOMPATIBLE WITH RFC[23]
    Illegal PstI site found at 1998
    Illegal PstI site found at 2591
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal PstI site found at 1998
    Illegal PstI site found at 2591
    Illegal AgeI site found at 1857
    Illegal AgeI site found at 4356
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 2430
    Illegal BsaI.rc site found at 864