IGEM:IMPERIAL/2006/project/popsblocker

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==Cre-LoxP System==
==Cre-LoxP System==
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==Motivations==
The use of Cre recombinase to excise regions of DNA between two loxP sites could be incorporated into part [http://parts.mit.edu/registry/index.php?title=Part:BBa_J37015 J37015]. Our aim is to insert two loxP sites into this part, which, when in place, would block the synthesis of any genes downstream of it, and when removed would allow their synthesis. Thus the Cre-loxP system would effectively act as a switch controlled by the activity of the promoter in front of the Cre enzyme.   
The use of Cre recombinase to excise regions of DNA between two loxP sites could be incorporated into part [http://parts.mit.edu/registry/index.php?title=Part:BBa_J37015 J37015]. Our aim is to insert two loxP sites into this part, which, when in place, would block the synthesis of any genes downstream of it, and when removed would allow their synthesis. Thus the Cre-loxP system would effectively act as a switch controlled by the activity of the promoter in front of the Cre enzyme.   
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==Part interface==
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(describe your black-box approach, define properties of inputs/outputs)
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{| border="0" width="100% align="center"
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|-
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|
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{| border="1" cellpadding="15" width="100%"
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|-
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! style="background:#66CC99"| INPUTS !!style="background:#FF3333"| OUTPUTS
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|-
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| input 1 || output 1
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|-
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| input 2 ||rowspan = "2" | output 2
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|-
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| input 3
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|}
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||
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[[Image:iGEM_IMPERIAL_Methodology_Specifications.png|right|350px]]
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|}
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(don't forget to insert a relevant image of your black box.)
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==Expected behavior and performances==
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(try to write your specifications in tables. Here are just 2 examples)
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{| border="1" width="100%"
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Interface specifiactions
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|-
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! style="background:lightgrey"|Characteristics !! Input 1 !! Input 2 !! Input 3 !! Output 1 !! Output 2
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|-
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| Range || [X1,Y1] || [X2,Y2] || [X3,Y3] || [X4,Y4] || [X5,Y5]
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|-
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| Tolerance || T1 ||T2 || T3 || T4|| T5
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|-
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| Transfer function ||  n/a  ||  n/a    ||  n/a  ||  insert image    || insert image
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|-
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| Latency||      ||      ||      ||        ||
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|}
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{| border="1" width="100%"
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System level specifications
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|-
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! style="background:lightgrey" width="100pt" |Properties !! Comments
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|-
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| Range||
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|-
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| Transfer function ||
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|-
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| Latency ||
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|-
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| Robustness || 
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|-
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| Variability  || 
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|-
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| properties 3 || 
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|-
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| properties 4 || 
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|}
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==Open issues==
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*list known issues
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*Using Cre recombinase as a switch appears to be, in theory, a very efficient and solid method.  
*Using Cre recombinase as a switch appears to be, in theory, a very efficient and solid method.  

Revision as of 12:26, 22 October 2006


Use this template

Contents

Cre-LoxP System

Motivations

The use of Cre recombinase to excise regions of DNA between two loxP sites could be incorporated into part J37015. Our aim is to insert two loxP sites into this part, which, when in place, would block the synthesis of any genes downstream of it, and when removed would allow their synthesis. Thus the Cre-loxP system would effectively act as a switch controlled by the activity of the promoter in front of the Cre enzyme.

Part interface

(describe your black-box approach, define properties of inputs/outputs)

INPUTS OUTPUTS
input 1 output 1
input 2 output 2
input 3

(don't forget to insert a relevant image of your black box.)

Expected behavior and performances

(try to write your specifications in tables. Here are just 2 examples)

Interface specifiactions
Characteristics Input 1 Input 2 Input 3 Output 1 Output 2
Range [X1,Y1] [X2,Y2] [X3,Y3] [X4,Y4] [X5,Y5]
Tolerance T1 T2 T3 T4 T5
Transfer function n/a n/a n/a insert image insert image
Latency


System level specifications
Properties Comments
Range
Transfer function
Latency
Robustness
Variability
properties 3
properties 4

Open issues

  • list known issues



  • Using Cre recombinase as a switch appears to be, in theory, a very efficient and solid method.
  • The parts are not available in the registry however, which means that we would have to make them.
  • The loxP sites are around 20 bases in length and so sequences can be be worked out and bought fairly easily.
  • The Cre recombinase site is around a kilobase in length. However we have been able to obtain the enzyme from one of the labs, and as it is isolated in a plasmid, this will make it much simpler for us to incorporate it into our design.
  • The Cre recombinase will also need a promoter - perhaps we could use one such as lacI? (we need to find out whether IPTG has any effect on other promoters though)
  • With all the parts available, we estimate that the length of time for implementation should be under a week.
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