Composite

Part:BBa_K1898250:Design

Designed by: Fiona Tsai   Group: iGEM16_TAS_Taipei   (2016-10-10)


Strong promoter + Strong RBS + GSR + 10x Histidine tag + Double terminator


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 7
    Illegal NheI site found at 30
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BamHI site found at 804
    Illegal BamHI site found at 1488
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 89
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal SapI site found at 1199
    Illegal SapI.rc site found at 1607


Design Notes

We did five silent point mutations to the GSR sequence to remove the two Pst1 and three EcoR1 internal cutting sites. The sequence was sent out for mutagenesis.

Primers were designed to remove the stop codon from GSR and to move the cDNA into iGEM Biobrick:

forward: 5' ATATgAATTCgCggCCg 3' (17)

reverse: 5' ATATCTgCAgCggCC 3' (15)

Our assembly method BioBrick RFC[10] shifted the open reading frame and created a stop codon in between GSR and 10x histidine tag. This is showed in this part's main page under sequencing. The sequence was sent to mutagenesis to add two base pairs in between the two genes.

Source

the cDNA of GSR was ordered from OriGene.

Primers were synthesized by Tri-I and mutagensis were performed by MissionBiotech.