Team:Harvard/Parts/LacI
From 2008.igem.org
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==Induction Test for LacI System with GFP== | ==Induction Test for LacI System with GFP== | ||
- | In preparing to make the lacI system for mtrB, we tested the IPTG inducibility of [http://partsregistry.org/wiki/index.php?title=Part:BBa_K098982 BBa_K098982] in ''E. coli''. In this system, the repressor | + | In preparing to make the lacI system for mtrB, we tested the IPTG inducibility of [http://partsregistry.org/wiki/index.php?title=Part:BBa_K098982 BBa_K098982] in ''E. coli''. In this system, the repressor is driven by a weak promoter. |
===Method=== | ===Method=== | ||
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===Results=== | ===Results=== | ||
- | Induction of GFP expression was observed at both 2 and 4 hours after adding IPTG. Levels of GFP expression in uninduced samples, however, remained relatively the same throughout the 4 hours. Meanwhile, IPTG induction was not observed in either the negative control ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K098981 BBa_K098981]) or the constitutive GFP control ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K098991 BBa_K098991]). | + | Induction of GFP expression was observed at both 2 and 4 hours after adding IPTG. Levels of GFP expression in uninduced samples, however, remained relatively the same throughout the 4 hours. Meanwhile, IPTG induction was not observed in either the negative control ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K098981 BBa_K098981]) or the constitutive GFP control ([http://partsregistry.org/wiki/index.php?title=Part:BBa_K098991 BBa_K098991]). Note that in the graph below, it appears that such an induction method is not reliable at 6 hours. |
+ | <div style="text-indent:0pt;color:black">[[Image:Baseline-corrected_of_lac_system.png|720px|thumb|center|IPTG successfully induces higher levels of GFP expression in cells containing [http://partsregistry.org/wiki/index.php?title=Part:BBa_K098982 BBa_K098982]]]</div> | ||
- | <div style="text-indent:0pt;color:black">[[Image:Lac.jpg| | + | |
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+ | Additionally, it appears that even the uninduced state of such a system has significant expression of GFP. Tighter control may require higher levels of lacI expression. This leakiness may also explain the difficulty we had in cloning mtrB (a gene toxic to ''E. coli''). | ||
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+ | <div style="text-indent:0pt;color:black">[[Image:Lac.jpg|650px|thumb|center|IPTG induction raw data]]</div> | ||
Revision as of 02:47, 30 October 2008
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