Composite
final2

Part:BBa_K2717021:Design

Designed by: Chenyu Liu   Group: iGEM18_BNU-China   (2018-10-09)


final2(GFP-hairpin-pchBA-terminator-emrr-RBS2000-GDH-v5 tag-his tag)


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1070
    Illegal NgoMIV site found at 1474
    Illegal NgoMIV site found at 1599
    Illegal NgoMIV site found at 1610
    Illegal NgoMIV site found at 1891
    Illegal NgoMIV site found at 2359
    Illegal AgeI site found at 4584
    Illegal AgeI site found at 5476
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 5458
    Illegal BsaI.rc site found at 644


Design Notes

GFP, pchBA, emrR binding promoter, emrR gene and gdh gene were obtained from plasmids or genomes by PCR methods. At the same time, V5tag and His tag were added after gdh to facilitate post-purification of proteins and westen bolt verification experiments. Then GFP and pchBA was fused by overlap to form a stretch, and the vector pUCyder was linearized by restriction enzyme linearization, next the gene fragment was ligated to the linearized vector by infusion.


Source

GFP is derived from the plasmid PSB1C3. pchBA is derived from the official kit. emrR binding promoter, emrR gene and gdh gene are all from the E.coli K12 genome.


References