http://2012.igem.org/wiki/index.php?title=Team:Cambridge/Ratiometrica/Parts&feed=atom&action=historyTeam:Cambridge/Ratiometrica/Parts - Revision history2024-03-28T09:11:39ZRevision history for this page on the wikiMediaWiki 1.16.0http://2012.igem.org/wiki/index.php?title=Team:Cambridge/Ratiometrica/Parts&diff=280287&oldid=prevEmmyft: Created page with "{{Template:Team:Cambridge/CAM_2012_TEMPLATE_HEADNEW}} <html> <script type="text/javascript"> $(document).keydown(function(e){ //e.which is set by jQuery for those browsers t..."2012-10-26T00:00:55Z<p>Created page with "{{Template:Team:Cambridge/CAM_2012_TEMPLATE_HEADNEW}} <html> <script type="text/javascript"> $(document).keydown(function(e){ //e.which is set by jQuery for those browsers t..."</p>
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= Ratiometrica Parts =<br />
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'''[http://partsregistry.org/wiki/index.php?title=Part:BBa_K911004<span style="color:#00000CD"><font size="4">Synthesised Ratiometric Luciferase construct in non-standard plasmid (Part:BBa_K911004) </font>]'''<br />
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This part was designed as a ratiometric luciferase reporter. The first promoter, hyperSpank, is LacI - repressed and controls the transcription of a (vibrio harveyi) luxA gene that has been fused at the N-terminus to an mOrange gene via a flexible linker. This was described by Dachuan Ke and Shiao-Chun Tu (2011) as having an additional peak in its emission spectrum at 560 nm, whereas the normal peak is at 490 nm. This is terminated by b0015. Downstream, pVEG controls the translation of the entire normal lux operon, which is again terminated by b0015.<br />
The idea is that the normal luciferase output acts as an internal control signal, to which the output of the induced luciferase with the spectral shift can be normalised. We designed this to be compatible with our cheap open-source sensing hardware.<br />
This part has some toxicity issues preventing it from being assembled in pSB1C3. We contacted iGEM HQ and were granted exemption from the pSB1C3 standard for this part.<br />
<br />
'''[http://partsregistry.org/wiki/index.php?title=Part:BBa_K911009<span style="color:#00000CD"><font size="4">Fluorescent ratiometric construct for standardizing promoter output(Part:BBa_K911009) </font>]'''<br />
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Biosensors may give unreliable outputs. This is due to differences in the number and state of the cells from test to test. By including an internal control signal, to which another inducible signal may be normalised, the reliability and reproducibility of a sensor may be significantly improved.<br />
The construct that uses an inducible eCFP (E0020) and a constitutively expressed eYFP (E0030) under the control of the constitutive promoter Pveg (K143012). The construct is optimized in both E. coli and B. subtilis, through the use of B. subtilis ribosome binding sites GsiB (K143020) and SpoVG (K143021).<br />
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'''[http://partsregistry.org/wiki/index.php?title=Part:BBa_K911006<span style="color:#00000CD"><font size="4">Non-standard backbone for luciferase construct (Part:BBa_K911006) </font>]'''<br />
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This is the sequence of the non-standard backbone that our luciferase construct, BBa_k911004, was synthesised in. It could not be assembled in the high copy number standard psb1c3 backbone due to toxicity issues. Attempts to assemble the 9kb insert into psb3c5, a low copy number biobrick plasmid have so far been unsuccessful.<br />
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