IGEM:IMPERIAL/2009/Encapsulation/Parts/Module3

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Contents

Inducible promoter

Parts and sequence details

Trigger

Problem

As the encapsulated cell is impermeable/not very receptive to solutes, we need physical means of induction


Possible Solutions

This could be solved in the following ways:

  • light
  • temperature
  • nutrient deprivation.

For temperature, it could be regulated using the following mechanisms:

  • Temperature sensitive lambda cI repressor
  • RNA thermometer
  • Cold sensor hdbB


Our solution

Using the temperature sensitive lambda cI repressor

BBa_K098995 by Harvard 2008 (on our 2009 distribution plate)
http://partsregistry.org/Part:BBa_K098995

This is a thermosensitive cI (clts) inducible system driven by a strong promoter.

Image:K098995.bmp

For strong promoter J23114
Variant RFP= 256 au

BBa_R0051 is a standard lambda cI regulated promoter.

This system is leaky so we have decided to make our own or obtain a commercially available thermoinducicble system.


Restriction Enzymes-Methylase system

Restriction Enzymes

Dpn1

  • blunt cutter
  • recognises GATC

Taq1

  • staggered cutter
  • recognises TCGA


Methylase

Dam Methylase

Weak constitutive promoter

To maintain a basal level of methylation

Using BBa_J23103
variant RFP= 17 au
A weak promoter in the popular constitutive promoter family.
http://partsregistry.org/Part:BBa_J23103

Overall design

The schematic below shows the overall design of the M3 system:


Useful reading

| Harvard Thermoinducible cI system

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