Team:Cambridge/References/ProjectBioluminescence/Luciferase

From 2010.igem.org

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==Photinus Pyralis (Firefly Luciferase)==
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{{:Team:Cambridge/Templates/headerbar|colour=#96d446|title=Bioluminescence: Luciferases}}
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==EPIC luciferase==
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==Mutant luciferase (P. pyralis)==
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*[http://www.ncbi.nlm.nih.gov/nuccore/AB261988.1 Mutant Enhanced Photon Initiating Complex (EPIC) Luciferase (P.pyralis) Sequence]
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*[http://www.ncbi.nlm.nih.gov/pubmed/10529195 Site-directed mutagenesis of firefly luciferase active site amino acids: a proposed model for bioluminescence color]
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*[http://www.ncbi.nlm.nih.gov/pubmed/12044905 Improved practical usefulness of firefly luciferase by gene chimerization and random mutagenesis]
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==luxCDABE (Vibrio Fischeri/Harvyi)==
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==Other Insect luciferases==
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*For colours we could use the click beetle luciferase [http://partsregistry.org/wiki/index.php/Part:BBa_J70005 red] or [http://partsregistry.org/wiki/index.php/Part:BBa_J70006 green]. They're only at the 'planning' stage though.
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*[Team:Cambridge/VibrioHarvei *Vibrio Harveyi]
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==Bacterial Bioluminescence==
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Harvyi
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*[http://arjournals.annualreviews.org/doi/pdf/10.1146/annurev.mi.42.100188.001055?cookieSet=1 paper with hard numbers, pathways]
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*Gram-negative,bioluminescent, marine bacteria  in the genus vibrio
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*[http://www.ncbi.nlm.nih.gov/pmc/articles/PMC372803/pdf/microrev00032-0137.pdf The definitive summary paper]
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*rod-shaped, motile (via polar flagella),
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*[[Team:Cambridge/VibrioFischeri| Vibrio Fischeri]]
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*facultatively anaerobic
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*[[Team:Cambridge/VibrioHarvei| Vibrio Harveyi]]
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*halophilic (extremophile for high salt)
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*[[Team:Cambridge/Phosphoreum| Vibrio Phosphoreum]]
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*competent for both fermentative and respiratory metabolism.  
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*[[Team:Cambridge/Photorhabdus| Photorhabdus]]
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*grow between 4°C and 35°C.
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*[[Team:Cambridge/ProjectBioluminescence/Luciferase/WikiGeneticsLuxCDABE| Genetics of luxCDABE (adapted from wikipedia)]]
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*lacks a LuxI/R quorum sensing system(which fischeri uses), instead employs a hybrid quorum-sensing circuit, detecting its autoinducer via a membrane-bound histidine kinase and using a phosphorelay.
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*[http://2006.igem.org/Lux_operon| 2006 attempts at BioBricking the lux operon]
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*[[Team:Cambridge/ProjectBioluminescence/Luciferase/Notes| Notes]]
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*[http://partsregistry.org/Lux The lux wiki from parts registry]
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*[http://www.ncbi.nlm.nih.gov/nuccore/AF170104.1?report=graph&log$=seqview NCBI Lux operon sequence]
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*[http://partsregistry.org/Part:BBa_G10001 parts registry lux operon] -currently waiting on an email to see how useful/available this part is
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*[[Team:Cambridge/ProjectBioluminescence/Luciferase/IMPORTANT INFO | IMPORTANT INFO]]
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* The Photorhabdus luminescens [http://partsregistry.org/Part:BBa_J70008 luxCDABE operon] in the registry. It's not well characterised
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[[Image:Luxoperondiagram.jpg]]
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==Plasmid experiment==
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*[http://departments.kings.edu/biology/lux/bacterial.html Vibrio plasmid experiment] -possible source of lux operon
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[[Image:4449975.pdf | article detailing proceedure and plasmids]]
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*[http://departments.kings.edu/biology/lux/index.html] - Transformation protocol for students from basics - includes several plasmids
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*Plan
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**set plasmid 555 into a BioBrick
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**replace the promoter? sensitivity tuners?
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*references
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** [http://www.pnas.org/content/84/19/6639.short Overproduction and purification of the luxR gene product: Transcriptional activator of the Vibrio fischeri luminescence system (Kaplan & Greenburg 1987) ]
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** [http://jb.asm.org/cgi/content/abstract/172/7/3974 Critical regions of the Vibrio fischeri luxR protein defined by mutational analysis (Slock et al 1990)]
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==Transformation Experiment to test Firefly luciferase in registry==
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* Firefly luciferase is in the registry as [http://partsregistry.org/Part:BBa_I712019 BBa_I712019], and have been sent to us as part of the spring 2010 distribution.
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*We need to find a new vector with a bacterial ribosyme binding site, initiator, and a stop codon. Having searched via the catalog in the plasmids section, then in expression plasmids, we have selected a constitutive plasmid:
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** The plasmid selected is [http://www.partsregistry.org/Part:BBa_J13002 BBa_J13002], which is a TetR repressed POPS/RIPS generator, again part of the spring 2010 distribution - kit plate 1, well 13B, plasmid pSB1A2. It was proven to work by UMN iGEM 2009.
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==Wavelengths==
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548 nm http://pubs.acs.org/doi/pdf/10.1021/bi7015052 <html><img src="http://www4a.wolframalpha.com/Calculate/MSP/MSP545519beai43119h6bi900003450dhf2577a40b8?MSPStoreType=image/gif&s=62&w=99&h=20"></html>
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613 nm http://www.promega.com/pnotes/85/10904_11/10904_11.pdf <html><img src="http://www4a.wolframalpha.com/Calculate/MSP/MSP52619bedhd64d9hi9240000123ei80gh721227b?MSPStoreType=image/gif&s=40&w=99&h=20"></html>
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<html>
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</div>
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</html>
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{| class="wikitable"
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!Luciferase!!Colour(at pH7.8)!!λ<sub>max</sub>(nm)!!(pH 7.8)!!(pH 6.0)!!Base change!!Amino acid change!!
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|-
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|Genjii||yellow-green||||562||609
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|-
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|C-M-1||orange||||607||614||G857-->A||Ser286-->Asn
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|-
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|C-M-2||red||||609||611||G976-->A||Gly326-->Ser
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|-
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|C-M-3||red||||612||612||C1297-->T||His433-->Tyr
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|-
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|C-M-4||yellow-orange||||595||609||C1354-->T||Pro452-->Ser
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|-
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|C-M-6||green||||558||558||G715-->A||Val239-->Ile
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|-
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|C-M-11||yellow||||565||612
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|}
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Latest revision as of 12:18, 7 October 2010