Difference between revisions of "Part:BBa K1132001:Design"
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This AND gate was built with one promoter-terminator couple surrounded by the Bxb1 integrase sites and a second terminator surrounded by the Tp901.1 integrase sites. This system is designed to be activated only in the presence of both recombinases (transcription of the output gene). The switch is permanent.<br><br>
 
This AND gate was built with one promoter-terminator couple surrounded by the Bxb1 integrase sites and a second terminator surrounded by the Tp901.1 integrase sites. This system is designed to be activated only in the presence of both recombinases (transcription of the output gene). The switch is permanent.<br><br>
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https://static.igem.org/mediawiki/2013/f/f8/2001.png<br><br>
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https://static.igem.org/mediawiki/2013/f/f8/2001.png           . . . . . .  https://static.igem.org/mediawiki/2013/4/43/2001_tab.png
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+
−
https://static.igem.org/mediawiki/2013/4/43/2001_tab.png
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<br><br>
 
<br><br>
  
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The input signals for this gate are the production of either one or both integrases Bxb1 and Tp901.1. The output can be choosen at will by assembling this biobrick to any ORF containing an RBS site. We also designed a test Biobrick of the gate (BBa_K1132031) with the RFP protein as output.
+
The input signals for this gate are the production of either one or both integrases Bxb1 and Tp901.1. The output can be choosen at will by assembling this biobrick to any ORF containing an RBS site. We also designed a test Biobrick of the gate ( [https://parts.igem.org/Part:BBa_K1132031 BBa_K1132031]) with the RFP protein as output.
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<br><br> This gate can be used in any regulation system, provided that the recombinases are assembled following the promoter of your choice with your specific regulations requirements. For example, if you want to activate the gate in presence of aTc and AHL, you just have to put the recombinase after the promoter activated by LuxR/AHL (BBa_R0065) and the promoter activated by aTc under the repression of TetR (BBa_R0040).
+
<br><br> This gate can be used in any regulation system, provided that the recombinases are assembled following the promoter of your choice with your specific regulations requirements. For example, if you want to activate the gate in presence of aTc and AHL, you just have to put the recombinase after the promoter activated by LuxR/AHL (BBa_R0065) and the promoter activated by aTc under the repression of TetR ([https://parts.igem.org/Part:BBa_R0040 BBa_R0040]).
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<br><br> Even a relatively small amount of recombinases can switch the DNA fragments. Therefore, it is really important to control the recombinases expression with a well-locked promoter. You can look at our specially designed regulation sequence (riboregulator) to get as low as possible any undesired expression and production of the recombinases (BBa_K1132005, BBa_K1132006, BBa_K1132007, BBa_K1132008, BBa_K1132042).
+
<br><br> Even a relatively small amount of recombinases can switch the DNA fragments. Therefore, it is really important to control the recombinases expression with a well-locked promoter. You can look at our specially designed regulation sequence (riboregulator) to get as low as possible any undesired expression and production of the recombinases ([https://parts.igem.org/Part:BBa_K1132005 BBa_K1132005], [https://parts.igem.org/Part:BBa_K1132006 BBa_K1132006], [https://parts.igem.org/Part:BBa_K1132007 BBa_K1132007], [https://parts.igem.org/Part:BBa_K11320308 BBa_K1132008], [https://parts.igem.org/Part:BBa_K1132042 BBa_K1132042]).
 
<br><br>In the present design, the strength of the promoter does not allow high level expression of the controlled output. However, change to stronger promoter than P7 should potentially lead to better expression levels.  
 
<br><br>In the present design, the strength of the promoter does not allow high level expression of the controlled output. However, change to stronger promoter than P7 should potentially lead to better expression levels.  
 
<br><br>Resetting the gate to its basal state requires a series of excisases capable of switching back the sequences to their native state.
 
<br><br>Resetting the gate to its basal state requires a series of excisases capable of switching back the sequences to their native state.
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<br>The same type of design was used to build a XOR gate (BBa_K1132002).
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<br>The same type of design was used to build a XOR gate ([https://parts.igem.org/Part:BBa_K1132002 BBa_K1132002]).
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+
  
  

Revision as of 11:07, 29 September 2013

AND gate with recombinases switching gene regulatory sequences


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    INCOMPATIBLE WITH RFC[12]
    Illegal NheI site found at 286
  • 21
    COMPATIBLE WITH RFC[21]
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 295
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 315
    Illegal BsaI.rc site found at 476


Design Notes

This AND gate was built with one promoter-terminator couple surrounded by the Bxb1 integrase sites and a second terminator surrounded by the Tp901.1 integrase sites. This system is designed to be activated only in the presence of both recombinases (transcription of the output gene). The switch is permanent.

2001.png . . . . . . 2001_tab.png

The input signals for this gate are the production of either one or both integrases Bxb1 and Tp901.1. The output can be choosen at will by assembling this biobrick to any ORF containing an RBS site. We also designed a test Biobrick of the gate ( BBa_K1132031) with the RFP protein as output.

This gate can be used in any regulation system, provided that the recombinases are assembled following the promoter of your choice with your specific regulations requirements. For example, if you want to activate the gate in presence of aTc and AHL, you just have to put the recombinase after the promoter activated by LuxR/AHL (BBa_R0065) and the promoter activated by aTc under the repression of TetR (BBa_R0040).

Even a relatively small amount of recombinases can switch the DNA fragments. Therefore, it is really important to control the recombinases expression with a well-locked promoter. You can look at our specially designed regulation sequence (riboregulator) to get as low as possible any undesired expression and production of the recombinases (BBa_K1132005, BBa_K1132006, BBa_K1132007, BBa_K1132008, BBa_K1132042).

In the present design, the strength of the promoter does not allow high level expression of the controlled output. However, change to stronger promoter than P7 should potentially lead to better expression levels.

Resetting the gate to its basal state requires a series of excisases capable of switching back the sequences to their native state.
The same type of design was used to build a XOR gate (BBa_K1132002).


Source

later

References

http://www.ncbi.nlm.nih.gov/pubmed/23539178
http://www.ncbi.nlm.nih.gov/nuccore/KC529327