Difference between revisions of "Part:BBa F2620"

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__NOTOC__
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<div style="font-family: Arial; padding: 00px; width: 680px; border: 0px solid #000000;">
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<partinfo>BBa_F2620 short</partinfo>
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{{Template:F2620 page header}}
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{{Template:F2620 Characterization Navbar}}
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<div style="padding: 00px; width: 680px; border: 0px solid #000000;">
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'''Description'''<br>
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A transcription factor, LuxR (''<partinfo>C0062</partinfo>'') that is active in the presence of cell-cell signaling molecule [[3OC6HSL|3OC<sub>6</sub>HSL]] is controlled by a TetR (''<partinfo>BBa_C0040</partinfo>'') regulated operator (''<partinfo>BBa_R0040</partinfo>'').  ''Device input'' is [[3OC6HSL|3OC<sub>6</sub>HSL]].  ''Device output'' is [[PoPS]] from a LuxR-regulated operator.  If used in a cell containing TetR then a second input signal such as [http://openwetware.org/wiki/ATc aTc] can be used to produce a logical AND function.  Get the pdf datasheet  [[:Image:EndyFig1-F2620DataSheet.pdf|here]].
  
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This device senses the 3OC<sub>6</sub>HSL concentration in the media and produces output PoPS according to the transfer function given below.  LuxR production is controlled by the tet repressible R0040.  Initial characterization work has been done on this device using T9002 as a test device.
 
  
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===Usage and Biology===
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'''Performance'''<br>
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<img src="'''Include File F2620Data'''"
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<center>
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{|{{Table}}
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!Experiment<sup>1</sup>
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!Characteristic<sup>1</sup>
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!Value<sup>1</sup>
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|-
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|rowspan="3"|[[Part:BBa_F2620:Transfer Function|'''Transfer Function''']]
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|''Maximum Output''
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|6.6 [[PoPS]] cell<sup>-1</sup>
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|-
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|''Hill coefficient''
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|1.6
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|-
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|[[Switch Point|''Switch Point'']]
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|1.5E-9 M [[3OC6HSL|3OC<sub>6</sub>HSL]], exogenous
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|-
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|[[Part:BBa_F2620:Response time|'''Response time:''']]
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|<1 min
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|-
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|rowspan="2"|[[Part:BBa_F2620:Specificity|'''Input compatibility''']]
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|''Strong response to''
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|[[3OC6HSL|3OC<sub>6</sub>HSL]], C<sub>6</sub>HSL , C<sub>7</sub>HSL, 3OC<sub>8</sub>HSL, C<sub>8</sub>HSL
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|-
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|''Weak response to''
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|C<sub>4</sub>HSL, C<sub>10</sub>HSL, C<sub>12</sub>HSL
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|-
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|rowspan="2"|[[Part:BBa_F2620:Stability|'''Stability''']]
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|[[Genetic Stability|''Genetic Stability'']]<br>(Low/High Input)
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|>92/>56 generations
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|-
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|[[Performance Stability|''Performance Stability'']]<br>(Low/High Input)
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|>92/>56 generations
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|-
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|rowspan="4"|Demand
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|rowspan="1"|Internal Demand<br>(Low/High Input)
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|Not measured
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|-
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|rowspan="2"|[[Transcription Demand|''Transcriptional output demand:'']]<br>(Low/High Input)<br>Nt = length of downstream transcript in nucleotides
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|(0/6xNt) nucleotides cell<sup>-1</sup> s<sup>-1</sup>
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|-
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|(0/1.5E-1xNt) RNAP cell<sup>-1</sup>
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|-
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|[[Growth Rate|''Growth Rate'']]<br>(Low/High Input)
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|54/59 min Doubling time
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|}
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</center>
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<sup>1</sup>Measured by Ania Labno and Barry Canton 2006-2007
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<div style="padding: 00px; width: 680px">
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'''Compatibility'''<br>
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[https://parts.igem.org/cgi/partsdb/pgroup.cgi?pgroup=cell ''Chassis:''] Device has been shown to work in ''<partinfo>BBa_V1000</partinfo>'',''<partinfo>BBa_V1001</partinfo>'',''<partinfo>BBa_V1002</partinfo>'', [[Chassis/Cell-Free_Systems/Commercial_E.coli_S30| E.Coli S30 Extract]] [[Chassis/Cell-Free_Systems/Commercial_E.coli_S30/F2620 | (data)]]<br>
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[[Plasmid backbones|''Plasmids:'']] Device has been shown to work on ''<partinfo>pSB3k3</partinfo>'' and ''<partinfo>pSB1A3</partinfo>''<br>
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[[Part Types|''Devices:'']] Device has been shown to work with ''<partinfo>BBa_E0430</partinfo>'', ''<partinfo>BBa_E0434</partinfo>'', ''<partinfo>BBa_E0240</partinfo>''<br>
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Crosstalk with systems containing ''<partinfo>BBa_C0040</partinfo>''.<br>
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[[Help:Signalling|''Cell-Cell Signaling Systems:'']] Crosstalk with devices using [[3OC6HSL|3OC<sub>6</sub>HSL]], C6HSL, C7HSL, C8HSL, C10HSL.
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</div>
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'''Contribution: Arizona_State 2016'''
  
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The transfer function indicates a switch point of approximately 2nM which was constant in repeated experiments (data not shown).
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Authors: Ernesto Luna, Brady Dennison, Cassandra Barrett, Jimmy Xu, Jiaqi Wu, Dr. Karmella Haynes
  
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The latency shows that fluorescent proteins appear within moments of the addition of AHL to the media. This suggests that PoPS output adjusts to a change in input very rapidly. This is expected given the current model of device operation.
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The 2016 Arizona_State team ran a series of induction tests on 10 AHL Senders (naturally-produced by an BL21 E.coli chassis) and 5 synthetic AHL variants using Bba_F2620 as the output device. This included a series of visual plate induction tests as well as an 8-hour GFP absorbance read. This contribution aims to expand the characterization of this part by identifying crosstalk partners as well as potential orthogonal systems.  
  
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<span class='h3bb'>Sequence and Features</span>
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The table below summarized the findings of our induction test. Note: these results are not conclusive and are simply what the collected data indicated from our 8-hour induction test.
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<partinfo>BBa_F2620 SequenceAndFeatures</partinfo>
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===Functional Parameters===
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<partinfo>BBa_F2620 parameters</partinfo>
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Get the device [http://openwetware.org/wiki/Image:F2620.de2-ania.ppt#filelinks datasheet]
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Two runs at two different concentrations (100nM and 1uM) using synthetic AHLs yielded some interesting differences, with synthetic p-Coumaroyl showing no induction while synthetic C12-HSL did induce. More in-depth figures and data can be found under the Experience tab of this part.  
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</div>

Latest revision as of 20:22, 11 October 2018

Input: 3OC6HSL
Output: PoPS

tetR
R0040		LuxR
I0462		lux pR
R0062
 BBa F2620Icon.png

Part Main Page        Transfer Function        Specificity        Response time        Stability        Add Data       


Description
A transcription factor, LuxR (BBa_C0062) that is active in the presence of cell-cell signaling molecule 3OC6HSL is controlled by a TetR (BBa_C0040) regulated operator (BBa_R0040). Device input is 3OC6HSL. Device output is PoPS from a LuxR-regulated operator. If used in a cell containing TetR then a second input signal such as [http://openwetware.org/wiki/ATc aTc] can be used to produce a logical AND function. Get the pdf datasheet here.


Performance

Experiment1 Characteristic1 Value1
Transfer Function Maximum Output 6.6 PoPS cell-1
Hill coefficient 1.6
Switch Point 1.5E-9 M 3OC6HSL, exogenous
Response time: <1 min
Input compatibility Strong response to 3OC6HSL, C6HSL , C7HSL, 3OC8HSL, C8HSL
Weak response to C4HSL, C10HSL, C12HSL
Stability Genetic Stability
(Low/High Input) 		>92/>56 generations
Performance Stability
(Low/High Input) 		>92/>56 generations
Demand Internal Demand
(Low/High Input) 		Not measured
Transcriptional output demand:
(Low/High Input)
Nt = length of downstream transcript in nucleotides 		(0/6xNt) nucleotides cell-1 s-1
(0/1.5E-1xNt) RNAP cell-1
Growth Rate
(Low/High Input) 		54/59 min Doubling time

1Measured by Ania Labno and Barry Canton 2006-2007

Compatibility
Chassis: Device has been shown to work in BBa_V1000,BBa_V1001,BBa_V1002, E.Coli S30 Extract (data)
Plasmids: Device has been shown to work on pSB3K3 and pSB1A3
Devices: Device has been shown to work with BBa_E0430, BBa_E0434, BBa_E0240
Crosstalk with systems containing BBa_C0040.
Cell-Cell Signaling Systems: Crosstalk with devices using 3OC6HSL, C6HSL, C7HSL, C8HSL, C10HSL.

Contribution: Arizona_State 2016

Authors: Ernesto Luna, Brady Dennison, Cassandra Barrett, Jimmy Xu, Jiaqi Wu, Dr. Karmella Haynes

The 2016 Arizona_State team ran a series of induction tests on 10 AHL Senders (naturally-produced by an BL21 E.coli chassis) and 5 synthetic AHL variants using Bba_F2620 as the output device. This included a series of visual plate induction tests as well as an 8-hour GFP absorbance read. This contribution aims to expand the characterization of this part by identifying crosstalk partners as well as potential orthogonal systems.

The table below summarized the findings of our induction test. Note: these results are not conclusive and are simply what the collected data indicated from our 8-hour induction test.


Two runs at two different concentrations (100nM and 1uM) using synthetic AHLs yielded some interesting differences, with synthetic p-Coumaroyl showing no induction while synthetic C12-HSL did induce. More in-depth figures and data can be found under the Experience tab of this part.