Composite

Part:BBa_K3505034

Designed by: Asteria Tsapadikou   Group: iGEM20_Thessaly   (2020-10-20)
Revision as of 17:09, 27 October 2020 by Asteriat (Talk | contribs) (→‎Usage and Biology)


pAndersonJ23115:tetO:RBS-lacI-terminator


Usage and Biology

This parts consists of AndersonJ23115 promoter BBa_K3505012 , LacI BBa_K3505003 and Double Terminator BBa_K3505017. By this way ,as AndersonJ23115 is a strong constitutive promoter, LacI is constitutively expressed. LacI binds to lac operator and inhibits the expression of a desirable transcription unit

Design Notes

The coding sequence was domesticated . We removed BsmBI ,BsaI , BtgZI, BpiI sites in order to be compatible with GoldenBraid and MoClo. The sequence is cloned in alpha vector BBa_K3505009 and has overhangs compatible for Golden Braid cloning.

Figure 2. The level A module of Tet-Off system : AndersonJ23115:TetO:RBS-LacI-Double terminator


Verification of Cloning

Fig.1:(U=Uncut C=Cut) Restriction digestion of TL: AndersonJ23115:Tet0-LacI-double terminator(C5-C8 ) with: HindIII + NheI (C5-C18) , Expected bands : 2573+891+427,Positive result : C5,C6

Sequence and Features


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 11
    Illegal NheI site found at 34
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    COMPATIBLE WITH RFC[25]
  • 1000
    COMPATIBLE WITH RFC[1000]


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Parameters
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