Reporter

Part:BBa_J04430

Designed by: Kristen DeCelle   Group: iGEM2005   (2005-06-09)

GFP coding device switched on by IPTG

Contains IPTG inducible promoter, an RBS, GFP (no LVA tag), and a terminator.


Usage and Biology

Isopropyl β-D-1-thiogalactopyranoside (IPTG) is a biological reagent which is a molecular mimic of allolactose and triggers trancrsiption of the lac operon and subsequent gene expression. The lac repressor, LacI, is also caused to dissociate from the promoter gene when lactose or allolactose-like module is present. LacI also binds to the major groove in the operator sequence and blocks T7 RNA polymerase from binding the promoter sequence. IPTG is therefore required in order to enable the binding and action of T7 RNA polymerase to transcribe the plasmid, or DNA fragment with T7 promoter. The IPTG promoter used in this system was Part BBa_R0010

The Ribosome binding site is used to initiate the translation of the protein coded for in the following coding sequence attracting ribosomes to the site of the start codon. The RBS in this system was Part BBa_B0034

The GFP was, the well characterised, Aequeora victoria jellyfish GFP Part BBa_E0040. This lacks the LVA degradation protein, allowing it to be characterised against a wider number of GFP containing systems.

There were two terminators used in this system; the T1 from E. coli rrnB and TE from coliphageT7.

N.B.The T1 from E.coli has a binding site for the VR primer therefore sequencing will be variably successful.


This part will be made using the standard ligation techniques.

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
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 870


Pictures

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[edit]
Categories
//classic/reporter/constitutive
Parameters
emissionGFP
excitation
tagNone