Difference between revisions of "Part:BBa K845000:Experience"

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In order to characterize the paraBAD promoter we used a transcriptional fusion of the promoter and gfpmut2 from Alon on pUA66 in BW25113 ΔcyaA.
 
In order to characterize the paraBAD promoter we used a transcriptional fusion of the promoter and gfpmut2 from Alon on pUA66 in BW25113 ΔcyaA.
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We followed the fluorescent expression of the GFP versus different concentrations of arabinose AND cAMP over time. We made this experiment in different growth media:
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We followed the fluorescent expression of the GFP versus different concentrations of arabinose AND cAMP over time. We made this experiment in different growth media (the different media did not affect bacterial growth):
 +
 
 
M9 complement with 0.03% of glucose and 0.03% of acetate (1)
 
M9 complement with 0.03% of glucose and 0.03% of acetate (1)
 +
 
M9 complement with 0.1% of glycerol (2)
 
M9 complement with 0.1% of glycerol (2)
 +
 
M9 complement with 0.1% of acetate (3)
 
M9 complement with 0.1% of acetate (3)
 +
  
 
See the protocol: http://2012.igem.org/Team:Grenoble/Biology/Protocols/AND_test
 
See the protocol: http://2012.igem.org/Team:Grenoble/Biology/Protocols/AND_test
  
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Those different media did not affect bacterial growth.
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Robustness test of the AND gate:
  
 
[[Image:Characterization_glucose.png]]
 
[[Image:Characterization_glucose.png]]
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[[Image:Characterization_glycerol.png]]
 
[[Image:Characterization_glycerol.png]]
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 +
As you can see on this figure even after few hours there is no fluorescent expression if one activator is missing.
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 +
 +
 +
With those different experiments we demonstrated that paraBAD works as expected. It will be activated only if there is arabinose and cAMP in the medium. It appears that both cAMP concentration and time response required for a half expression of GFP depends on the medium.
 +
 +
For more information: http://2012.igem.org/Team:Grenoble/Biology/AND_gate
  
 
===User Reviews===
 
===User Reviews===

Latest revision as of 15:25, 29 September 2012

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K845000

In order to characterize the paraBAD promoter we used a transcriptional fusion of the promoter and gfpmut2 from Alon on pUA66 in BW25113 ΔcyaA. We followed the fluorescent expression of the GFP versus different concentrations of arabinose AND cAMP over time. We made this experiment in different growth media (the different media did not affect bacterial growth):

M9 complement with 0.03% of glucose and 0.03% of acetate (1)

M9 complement with 0.1% of glycerol (2)

M9 complement with 0.1% of acetate (3)


See the protocol: http://2012.igem.org/Team:Grenoble/Biology/Protocols/AND_test

Robustness test of the AND gate:

Characterization glucose.png

Characterization acetate.png

Characterization glycerol.png

As you can see on this figure even after few hours there is no fluorescent expression if one activator is missing.


With those different experiments we demonstrated that paraBAD works as expected. It will be activated only if there is arabinose and cAMP in the medium. It appears that both cAMP concentration and time response required for a half expression of GFP depends on the medium.

For more information: http://2012.igem.org/Team:Grenoble/Biology/AND_gate

User Reviews

UNIQbd7727dcdbfe2ebf-partinfo-00000000-QINU UNIQbd7727dcdbfe2ebf-partinfo-00000001-QINU