Regulatory

Part:BBa_K3866003

Designed by: Ioanna Gkoni   Group: iGEM21_Thessaly   (2021-09-23)
Revision as of 08:26, 15 October 2021 by Igkoni (Talk | contribs) (→‎Usage and Biology)


Partial T5 Promoter - CuO- RBS GB compatible with A1-B2

Partial T5 promoter is a strong constitutive Promoter that can be used in the toxin-antitoxin system with the Cym repressor that our team uses in their system.

Fig.2:Partial T5 Promoter with Lactose Operator and Cumate Operator with compatible ends for Golden Braid Cloning

Usage and Biology

T5 promoter has been used for many years for protein expression in E.coli. Partial T5 promoter is used with the downstream Operator CuO.

Figure 6. Growth (OD600) of E. coli MC1061 cells transformed with N20-BglX (N20), native-peptide BglX (SP) or an empty vector (EMPTY), in a minimal M9 medium with Glucose and Cellobiose as a carbon source (2).

Design Notes

The sequence was domesticated . We removed BsmBI ,BsaI , BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is cloned in pUPD2 BBa_K3505007 and has overhangs compatible for GoldenBraid cloning. This has position A1-B2.

Fig.1:The overhangs of this part in the GoldenBraid Grammar


Sequence and Features


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]

Source

Synthesized by Twist Biosciences.

References

  • Choi, Y. J., Morel, L., Le François, T., Bourque, D., Bourget, L., Groleau, D., Massie, B., & Míguez, C. B. (2010). Novel, versatile, and tightly regulated expression system for Escherichia coli strains. Applied and environmental microbiology, 76(15), 5058–5066. https://doi.org/10.1128/AEM.00413-10
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