Composite
WT DNAP

Part:BBa_K2918022

Designed by: TUDelft2019   Group: iGEM19_TUDelft   (2019-09-27)
Revision as of 19:13, 8 December 2019 by Weronika77 (Talk | contribs)

WT T7 promoter - Universal RBS - Φ29 DNAP - WT T7 terminator

A level 1 MoClo construct with a Wild-Type T7 promoter, a universal RBS, phi29 DNAP and a T7 terminator.

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
    INCOMPATIBLE WITH RFC[25]
    Illegal AgeI site found at 1770
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 60
    Illegal BsaI.rc site found at 86

The construct is confirmed by sequencing and there are no mutations.

Toxicity

Our Sci-Phi 29 tool is based on four components of the Φ29 bacteriophage: DNAP, TP, p5 and p6. However, overexpression of these proteins are toxic for the cell. In order to determine the optimal expression levels of the proteins in live cells, we carried out viability assays in E. coli BL21(DE3) pLysS. The results are shown in the graphs below.

  • Figure 3A: Normalized maximum growth rate of phi29 DNAP under different promoter strengths (weak, medium, Wild-Type) with no IPTG induction
  • Figure 3B: Normalized maximum growth rate of phi29 DNAP under different promoter strengths (weak, medium, Wild-Type) with 1 mM IPTG induction
  • Figure 3C: Normalized maximum growth rate of phi29 DNAP under different promoter strengths (weak, medium, Wild-Type) with 10 mM IPTG induction


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Categories
Parameters
None