Difference between revisions of "Part:BBa R0077:Experience"
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| − | First, we induced generator cell with 10-3M IPTG for 16 hours, then centrifuged cells and collect the supernatant. We use this supernatant as original 3OH-C14:1-HSL solution，then we diluted it to 10-7, 5×10-7, 10-6, 5×10-6, 10-5, 6×10-5, 8×10-5, 10-4, 2×10-4, 4×10-4, 7×10-4, 10-3, 3×10-3, 6×10-3, 8×10-3, 10-2 of original solution concentration respectively. Using these diluted supernatants to induce receiver cells (OD600 0.4) at 37℃ for 3 hours, then centrifuged cells and resuspended in phosphate buffer solution (PBS). The GFP fluorescence of each culture was obtained using enzyme-labeled assay. The result is shown in figure | + | First, we induced generator cell with 10-3M IPTG for 16 hours, then centrifuged cells and collect the supernatant. We use this supernatant as original 3OH-C14:1-HSL solution，then we diluted it to 10-7, 5×10-7, 10-6, 5×10-6, 10-5, 6×10-5, 8×10-5, 10-4, 2×10-4, 4×10-4, 7×10-4, 10-3, 3×10-3, 6×10-3, 8×10-3, 10-2 of original solution concentration respectively. Using these diluted supernatants to induce receiver cells (OD600 0.4) at 37℃ for 3 hours, then centrifuged cells and resuspended in phosphate buffer solution (PBS). The GFP fluorescence of each culture was obtained using enzyme-labeled assay. The result is shown in figure 1. |
<center>[[Image:Refigure1.png]]</center> | <center>[[Image:Refigure1.png]]</center> | ||
| − | ''Figure | + | ''Figure 1. Dose Response Curve. Vertical axis RFU/OD represents GFP expression measured through enzyme-labeled assay and the Horizontal axis represent different relative dilution concentration of 3OH-C14:1-HSL. Data was fitted with Hill function. |
'' | '' | ||
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We induced E.coli with receiver plasmid using supernatant diluted 10-2 times, and measure OD and absorbance of GFP every 15 minutes. | We induced E.coli with receiver plasmid using supernatant diluted 10-2 times, and measure OD and absorbance of GFP every 15 minutes. | ||
<center>[[Image:LM9.png]]</center> | <center>[[Image:LM9.png]]</center> | ||
| − | ''Figure | + | ''Figure 2. Time response curve of cin receiver system. We measured RFU and OD600 every 15 minutes and experiment performed in 3 hours.'' |
Coordinatiblity | Coordinatiblity | ||
<center>[[Image:LM12a.png|600px]]</center> | <center>[[Image:LM12a.png|600px]]</center> | ||
| − | ''Cin receiver system cell distribution of GFP fluorescence intensity.'' | + | ''Figure 3. Cin receiver system cell distribution of GFP fluorescence intensity.'' |
Revision as of 23:50, 5 October 2011
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UNIQ018e5c4176d4f3bb-partinfo-00000000-QINU UNIQ018e5c4176d4f3bb-partinfo-00000001-QINU First, we induced generator cell with 10-3M IPTG for 16 hours, then centrifuged cells and collect the supernatant. We use this supernatant as original 3OH-C14:1-HSL solution，then we diluted it to 10-7, 5×10-7, 10-6, 5×10-6, 10-5, 6×10-5, 8×10-5, 10-4, 2×10-4, 4×10-4, 7×10-4, 10-3, 3×10-3, 6×10-3, 8×10-3, 10-2 of original solution concentration respectively. Using these diluted supernatants to induce receiver cells (OD600 0.4) at 37℃ for 3 hours, then centrifuged cells and resuspended in phosphate buffer solution (PBS). The GFP fluorescence of each culture was obtained using enzyme-labeled assay. The result is shown in figure 1.
Figure 1. Dose Response Curve. Vertical axis RFU/OD represents GFP expression measured through enzyme-labeled assay and the Horizontal axis represent different relative dilution concentration of 3OH-C14:1-HSL. Data was fitted with Hill function.
Time Dependence
We induced E.coli with receiver plasmid using supernatant diluted 10-2 times, and measure OD and absorbance of GFP every 15 minutes.
Figure 2. Time response curve of cin receiver system. We measured RFU and OD600 every 15 minutes and experiment performed in 3 hours.
Coordinatiblity
Figure 3. Cin receiver system cell distribution of GFP fluorescence intensity.