User:Isis Trenchard/Notebook/BioE44 Stuff/2010/02/26

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To do list grow and test color generator strains figure out how I'm going to do this look up Cambridge generator info figure out how color production was assayed start compiling some course materials start working on course website read about arsenic sensitive promoter Info on parts we have (taken from Cambridge iGEM wiki) Purple Generator Part Number Plasmid What is it BBaK274002 pJ241 (DNA 2.0 vector) Violacein. Produces a purple pigment (violacein) from L-tyrosine. The operon contains five genes (VioA-E) each with their own ribosome binding sites. Green Generators Part Number Plasmid What is it BBaK274004 pJ241 (DNA 2.0 vector) Vio operon ABCE. Produces a light green pigment from L-tyrosine. Formed from the vio operon biobrick (BBa_K274002) with the vioD gene removed by restriction digest with BglII and BclI. This sequence contains four genes, vioA, vioB vioC and vioE, each preceded by their own ribosome binding site. BBaK274003 pJ241 (DNA 2.0 vector) Vio operon ABDE. Produces a dark green pigment from L-tyrosine. Formed from the vio operon biobrick (BBa_K274002) with the vioC gene removed by restriction digest with BamHI and BglII. This sequence contains four genes, vioA, vioB vioD and vioE, each preceded by their own ribosome binding site. Red Generators Part Number Plasmid What is it BBaK274100 pSB1A2 CrtEBI with rbs. This Composite Biobrick is created by standard assembly of 3 basic Biobricks coding for enzymes CrtE, CrtB and CrtI (each with rbs). Together, enzymes CrtE, CrtB and CrtI convert colourless farnesyl pyrophosphate to red lycopene (via intermediates geranylgeranyl pyroiphosphate and phytoene). The red lycopene pigment can then be used as a coloured signal output, e.g. for biosensors. BBaK274110 pSB1A2 CrtEBI under constitutive promoter. This Biobrick is created by putting enzyme cassette CrtEBI (with individual rbs) of Part BBa_K274100 under constitutive promoter R0011. Amount of lycopene produced can be measured by photospectrometer with absorbance at 475nm (lycopene extraction using acetone). BBaK274120 pSB2K3 CrtEBI under pBad promoter. This Biobrick is created by putting enzyme cassette CrtEBI (with individual rbs) of Part BBa_K274100 under arabinose-induced promoter I0500. Amount of lycopene produced can be measured by photospectrometer with absorbance at 475nm (lycopene extraction using acetone). Orange Generators Part Number Plasmid What is it BBaK274200 pSB1A2 CrtEBIY with rbs. This Composite Biobrick is created by standard assembly of 4 basic Biobricks coding for enzymes CrtE, CrtB, CrtI and CrtY (each with rbs). Together, enzymes CrtE, CrtB, CrtI and CrtY convert colourless farnesyl pyrophosphate to orange beta-carotene (via intermediates geranylgeranyl pyroiphosphate, phytoene and lycopene). The orange beta-carotene pigment can then be used as a coloured signal output, e.g. for biosensors. BBaK274210 pSB1A2 CrtEBIY under constitutive promoter. This Biobrick is created by putting enzyme cassette CrtEBIY (with individual rbs) of Part BBa_K274200 under constitutive promoter R0011. Amount of beta-carotene produced can be measured by photospectrometer with absorbance at 455nm (beta-carotene extraction using acetone). BBaK274220 pSB2K3 CrtEBIY under pBad promoter. This Biobrick is created by putting enzyme cassette CrtEBIY (with individual rbs) of Part BBa_K274200 under arabinose-induced promoter I0500. Amount of beta-carotene produced can be measured by photospectrometer with absorbance at 455nm (beta-carotene extraction using acetone). Stuff that I should probably read Purple generator August PR, Grossman TH, Minor C, Draper MP, MacNeil IA, Pemberton JM, Call KM, Holt D, and Osburne MS. Sequence analysis and functional characterization of the violacein biosynthetic pathway from Chromobacterium violaceum. J Mol Microbiol Biotechnol. 2000 Oct;2(4):513-9. Antônio RV and Creczynski-Pasa TB. Genetic analysis of violacein biosynthesis by Chromobacterium violaceum. Genet Mol Res. 2004 Mar 31;3(1):85-91.