Difference between revisions of "Part:BBa K325100"

 
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<div style="font-family: Arial; padding: 00px; width: 680px; border: 0px solid #000000;">
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{{Template:K325219 page header}}
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{{Template:K325219 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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This part generates a mutant of the luciferase from the North American firefly (L.cruciata) as well as this species' luciferin regenerating enzyme (LRE). It is under the control of an Arabinose induced promoter (pBAD).
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D-Luciferin has to be added to obtain light output.
  
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__NOTOC__
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The part is codon optimised for expression in ''E.coli''.
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<partinfo>BBa_K325100 short</partinfo>
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This is a mutant luciferase that is very much brighter than wild-type.  It is supplied in an operon with an enzyme (LRE) that regenerates its substrate. When supplied with luciferin and placed under a promoter it will emit light at a green wavelength.  If D-cysteine is added the LRE will be able to regenerate the luciferin substrate.
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THIS PAGE IS CURRENTLY BEING UPDATED.  
  
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<!-- Add more about the biology of this part here
 
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===Usage and Biology===
 
  
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<!-- -->
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'''Performance'''<br>
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<span class='h3bb'>Sequence and Features</span>
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<center>
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<partinfo>BBa_K325100 SequenceAndFeatures</partinfo>
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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 the [http://2010.igem.org/Team:Cambridge Cambridge iGEM team 2010]
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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 ''Top 10 (Invitrogen)''<br>
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[[Plasmid backbones|''Plasmids:'']] Device has been shown to work on ''<partinfo>pSB1C3</partinfo>'' <br>
  
  
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<!-- Uncomment this to enable Functional Parameter display
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</div>
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===Functional Parameters===
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<partinfo>BBa_K325100 parameters</partinfo>
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'''References'''<br>
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<!-- -->
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[http://www.ncbi.nlm.nih.gov/pubmed/18949818 '''[1&#x5d;:'''] S.M. Marques and J.C.G. Esteves da Silva, (2009) Firefly Bioluminescence: A Mechanistic Approach of Luciferase Catalyzed Reactions,''Life'' '''61''', 6-17.
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</div>
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[http://www.nature.com/nature/journal/v440/n7082/abs/nature04542.html '''[2&#x5d;:'''] T. Nakatsu ''et al.'' (2006) Structural Basis for the spectral difference in luciferase bioluminescence, ''Nature'' '''440'''(16), 372-376.
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[http://www.ncbi.nlm.nih.gov/pubmed/11457857 '''[3&#x5d;:'''] K. Gomi and N. Kajiyama, (2001) Oxyluciferin, a Luminescence Product of Firefly Luciferase, Is Enzymatically Regenerated into Luciferin, ''The Journal of Biological Chemistry'', '''276'''(39), 36508-36513.

Revision as of 21:45, 23 October 2010

Input: L-Arabinose
Output: Light

pBad/araC
I0500		Luciferase/LRE
K325210
 Cambridge-Eglowli.png

Part Main Page        Arabinose -> Light        Add Data       


Description
This part generates a mutant of the luciferase from the North American firefly (L.cruciata) as well as this species' luciferin regenerating enzyme (LRE). It is under the control of an Arabinose induced promoter (pBAD). D-Luciferin has to be added to obtain light output.

The part is codon optimised for expression in E.coli.

THIS PAGE IS CURRENTLY BEING UPDATED.


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 the [http://2010.igem.org/Team:Cambridge Cambridge iGEM team 2010]

Compatibility
Chassis: Device has been shown to work in Top 10 (Invitrogen)
Plasmids: Device has been shown to work on pSB1C3


References
[http://www.ncbi.nlm.nih.gov/pubmed/18949818 [1]:] S.M. Marques and J.C.G. Esteves da Silva, (2009) Firefly Bioluminescence: A Mechanistic Approach of Luciferase Catalyzed Reactions,Life 61, 6-17.

[http://www.nature.com/nature/journal/v440/n7082/abs/nature04542.html [2]:] T. Nakatsu et al. (2006) Structural Basis for the spectral difference in luciferase bioluminescence, Nature 440(16), 372-376.

[http://www.ncbi.nlm.nih.gov/pubmed/11457857 [3]:] K. Gomi and N. Kajiyama, (2001) Oxyluciferin, a Luminescence Product of Firefly Luciferase, Is Enzymatically Regenerated into Luciferin, The Journal of Biological Chemistry, 276(39), 36508-36513.