Source:
Generated By: https://synbiohub.org/public/igem/igem2sbol/1
Created by: Siming Ma
Date created: 2009-09-17 11:00:00
Date modified: 2015-05-08 01:11:45
CrtEBIY with rbs
| Types | DnaRegion |
| Roles | engineered_region Composite |
| Sequences | BBa_K274200_sequence (Version 1) |
Description
This Composite Biobrick is created by standard assembly of Part BBa_K118014 (CrtE with rbs), Part BBa_K118006 (CrtB with rbs), Part BBa_K118005 (CrtI with rbs) and Part BBa_K118013 (CrtY with rbs), which are submitted by previous iGEM teams. The whole cassette is on plasmid pSB1A2 (high copy, Ampicillin resistance).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.
There is already individual ribosome binding site before each enzyme gene sequence. Internal restriction sites have been removed by previous iGEM teams. Please refer to Parts BBa_K118014, BBa_K118006, BBa_K118005 and BBa_K118013 for more information on individual Biobricks components.
This Composite Biobrick has been put under constitutive promoter R0011 (see Part BBa_K274210) and arabinose-inducible promoter I0500 (see Part BBa_274220), transformed and tested in E.coli strain MG1655.
Amount of beta-carotene produced can be measured by photospectrometer with absorbance at 455nm (beta-carotene extraction using acetone).
A related Composite Biobrick is Part BBa_K274100 (CrtEBI with rbs), which "stops" at one step before beta-carotene, producing (and retaining) red lycopene pigment.
Notes
The colourless precursor, farnesyl pyrophosphate, is naturally produced in E.coli using Non-mevalonate Pathway (from pyruvate and glyceraldehyde-3-phosphate).Our experiences showed that this Biobrick works better in E.coli strains with up-regulated level of precursors. E.coli strain MG1655 demonostrates higher level of pigment (beta-carotene) production than strain TOP10.
Source
Gene coding sequences come from Pantoea ananatis (formerly Erwinia uredovora) (Accession number D90087), a type of Gram negative Enterobacteriaceae.This Composite Biobrick is created by standard assembly of Part BBa_K118014, Part BBa_K118006, Part BBa_K118005 and Part BBa_K118013.
| Access | Instance | Definition |
| public public public public public public public public | BBa_K118012 BBa_I742151 BBa_J15001 BBa_K118002 BBa_J15001 BBa_K118003 BBa_K118012 BBa_K118008 | BBa_K118012 BBa_I742151 BBa_J15001 BBa_K118002 BBa_J15001 BBa_K118003 BBa_K118012 BBa_K118008 |