Regulatory

Part:BBa_K2273111:Design

Designed by: Nina Lautenschlaeger   Group: iGEM17_TU_Dresden   (2017-10-02)
Revision as of 17:17, 30 October 2017 by NinaL (Talk | contribs) (→‎References)

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PblaZ Promoter controlling gene expression of blaZ in S. aureus


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
    COMPATIBLE WITH RFC[1000]


Design Notes

This Promoter was designed to be cloned in front of the luxABCDE operon in the pBS3C vector backbone and therefore the RFC10 Standard Restriction sites have been added as prefix and suffix.

Prefix with EcoRI, NotI, XbaI GAATTCGCGGCCGCTTCTAGA
Suffix with SpeI, NotI and PstI ACTAGTAGCGGCCGCTGCAGA

Sites of restriction enzymes generating compatible overhangs are indicated by sharing one color. (EcoRI and PstI are marked in blue, NotI in green, XbaI and SpeI in red

As the overall promoter sequence was quite short, we created it by ordering to long Primers that have a 20 base pair overlap:

Forward Primer Sequence containing RFC10 prefix: 5`-GATCGAATTCGCGGCCGCTTCTAGATTCAAATATTTATAATAAACAATTGACATCAATATTACAATTGTAATATTATTG-3`

Revers Primer Sequence containing RFC10 suffix: 5`-GATCCTGCAGCGGCCGCTACTAGTATATTACAGTTGTAATTTTTATAAATCAATAATATTACAATTGTAATATTGATG-3`



Source

This part sequence was derived from the genome sequence of Staphylococcus aureus N315 found in the NCBI database.

References

1Leticia I. Llarrull, Mary Prorok, and Shahriar Mobashery (2010) Binding of the Gene Repressor BlaI to the bla Operon in Methicillin-Resistant Staphylococcus aureus. Biochemistry 49(37), 7975–7977