Difference between revisions of "Part:BBa K395301:Experience"
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[[Image:Tokyotech_ompc_graph.jpg|thumb|center|400px|Figure 1. Induction of new'' OmpC'' series in high osmolarity medium ]] | [[Image:Tokyotech_ompc_graph.jpg|thumb|center|400px|Figure 1. Induction of new'' OmpC'' series in high osmolarity medium ]] | ||
− | + | In order to characterize P''ompC(C)'' [ https://parts.igem.org/wiki/index.php?title=Part:BBa_K395301 BBa_395301], P''ompC(CB)'' [https://parts.igem.org/wiki/index.php?title=Part:BBa_K395302 BBa395302] , P''ompC(CS1)'' [https://parts.igem.org/wiki/index.php?title=Part:BBa_K395303 BBa_395303], each promoter was attached to GFP and its transcriptional activity was measured through the GFP expression. <br> | |
− | P''ompC(C)''[https://parts.igem.org/wiki/index.php?title=Part:BBa_K395301 | + | PompC(C)-GFP, PompC(CB)-GFP PompC(CS1)-GFP and PompC(WT)-GFP on pSB3K3 was introduced into E. coli strain MG1655. A strain containing placI<sup>q</sup>-GFP, plasmid (BBa_J54202), a constitutive GFP expressive promoter, and promoterless GFP reporter plasmid (BBa_J54103) was used as a positive control and negative control respectively. <br><br> |
Overnight cultures of reporter strains grown at 37 °C in Medium A containing appropriated antibiotics were diluted at least 1:100 in the medium and incubated at 37 °C as fresh cultures. After their OD590 reached 0.2, the fresh culture was diluted 1 : 3 into 2 ml of pre-warmed medium A. For high osmolarity conditions, the cultures were diluted with sucrose supplemented medium to the final concentration of 15% (wt/vol). After 4 hours of induction, fluorescence intensity was measured with fluorometer. <br><br> | Overnight cultures of reporter strains grown at 37 °C in Medium A containing appropriated antibiotics were diluted at least 1:100 in the medium and incubated at 37 °C as fresh cultures. After their OD590 reached 0.2, the fresh culture was diluted 1 : 3 into 2 ml of pre-warmed medium A. For high osmolarity conditions, the cultures were diluted with sucrose supplemented medium to the final concentration of 15% (wt/vol). After 4 hours of induction, fluorescence intensity was measured with fluorometer. <br><br> | ||
After 4 hours of sucrose induction, transcriptional activity of P''ompC(CB)''-GFP and P''ompC(CS1)''-GFP increased 2.5 folds and 2.3 folds respectively. However, significant amount of leaky expression was found in P''ompC(CS1)''-GFP without induction. In contrast, under the same conditions, we found no significant difference of GFP expression in P''ompC(C)''-GFP and P''ompC(WT)''-GFP. [http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC ...see more about P''ompC'' series] | After 4 hours of sucrose induction, transcriptional activity of P''ompC(CB)''-GFP and P''ompC(CS1)''-GFP increased 2.5 folds and 2.3 folds respectively. However, significant amount of leaky expression was found in P''ompC(CS1)''-GFP without induction. In contrast, under the same conditions, we found no significant difference of GFP expression in P''ompC(C)''-GFP and P''ompC(WT)''-GFP. [http://2010.igem.org/Team:Tokyo_Tech/Project/wolf_coli/New_Series_of_PompC ...see more about P''ompC'' series] |
Revision as of 12:37, 26 October 2010
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Tokyo Tech iGEM2010 |
In order to characterize PompC(C) [ https://parts.igem.org/wiki/index.php?title=Part:BBa_K395301 BBa_395301], PompC(CB) BBa395302 , PompC(CS1) BBa_395303, each promoter was attached to GFP and its transcriptional activity was measured through the GFP expression. |
UNIQ07f7a24380a58106-partinfo-00000001-QINU