Team:Cambridge/References/ProjectBioluminescence

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=Bioluminescence=
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{{:Team:Cambridge/Templates/headerbar|colour=#96d446|title=Bioluminescence: Introduction}}
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==The Plan==
==The Plan==
* Characterise the transfer function of PoPs + CHBT  --> Light
* Characterise the transfer function of PoPs + CHBT  --> Light
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* Our Characterisation should look like [http://www.nature.com/nbt/journal/v26/n7/abs/nbt1413.html this].
==The Theory==
==The Theory==
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[[Image:Cam-luci-cycle.jpg | 600px]]
[[Image:Cam-luci-cycle.jpg | 600px]]
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==Increasing Light Emission==
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==Amazing mutant==
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*[http://www.ncbi.nlm.nih.gov/pubmed Paper on isolation of H-LRE and G-LRE]
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paper on cloning the lux operon from Ponyfish Photobacterium leiognathi into E.coli, by the E.coli mutant 43R the luminescence output could be increased dramatically to near native levels. --> find out what 43R does! [http://www3.interscience.wiley.com/cgi-bin/fulltext/120766166/PDFSTART Chan et al. 1991] suspects negative effectors in wild-type E.coli, but not in the mutant or in P.leiognathi, inhibit luminescence. If this is the case, it might be difficult to mimic the mutant phenotype with genes imported on a plasmid.
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Previous attempts to improve light production of luciferase reaction
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Pyruvate orthophosphate dikinase (PPDK) converts AMP + PPi to ATP (AMP + PPi + Phosphoenol Pyruvate -> ATP + Pyruvate + Phosphate) => light emission intensified in presence of low ATP concentration
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[http://www.springerlink.com/content/vlxb2c59xut1htd8/fulltext.pdf Ulitzur et al. 1997] describes phenotypes like 43R being generated by deletions in the H-NS gene, fancy that. Apparently H-NS acts as a pleiotropic transcriptional repressor, with particularly strong effects on a certain set of promoters. Mutants have a reduced expression of a flagellar protein and are non-motile, but fully viable. In fact they appear to grow faster. However, H-NS has been implicated in E.coli thermostability.
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*[http://www.ncbi.nlm.nih.gov/pubmed/1332531 Enhanced luciferase activity through cytodine nucleotides]
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*[http://www.ncbi.nlm.nih.gov/pubmed/12044905 Gene chimerisation to improve practical usefulness of firefly luciferase]
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*[http://www.ncbi.nlm.nih.gov/pubmed/17540326 Genetically modified firefly luciferase - the EPIC luciferase]
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==Sequences==
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==ADLA==
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*[http://www.ncbi.nlm.nih.gov/nucleotide/14331151?report=genbank&log$=nucltop&blast_rank=1&RID=4EUAXMJW01R A-LRE (P.pyralis) Sequence]
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The Abrupt Decline in Luciferase Activity is a phenomenon described for bacterial luciferase in E.coli in stationary phase of growth (see [http://www.springerlink.com/content/w73k840k27866462/fulltext.pdf Koga et al. 2005]) The paper identifies two mutants, HupA and an N-terminal deletion of H-NS for which ADLA does not occur. hns205, the mutation necessary for quiescence is a C-terminal deletion. We should find out whether the phenotypes are the same, which would be sweet if we ended up doing quiescence as well.
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*[http://www.ncbi.nlm.nih.gov/nuccore/AB261988.1 Mutant Luciferase (P.pyralis) Sequence]
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Latest revision as of 12:16, 7 October 2010