Difference between revisions of "Part:BBa K1139150:Experience"

Revision as of 05:16, 27 September 2013

Prm/lac-GFP-TT

2. Assay protocol
1. Prepare overnight culture of each cell at 37°C for 12 hours.
2. Take 30 µL of the overnight cultures into LB (3 mL) containing antibiotics (Amp 50 µg/mL) and 1 mM of IPTG (=> fresh culture)
We added IPTG in order to make sure to repress the expression of LacI derived from E. coli genome.
3. After 4 hours of induction, flow cytometer measurements for GFP expression.

Results

Fig. 1 shows the fluorescence intensity detected by flow cytometer.
Fig. 2 is the extracted data which shows the comparison of N99 (IPTG+, with constitutive CI) and JM2.300 (IPTG+, without CI).

Fig. 1. Fluorescence intensity detected by flow cytometer

Fig. 2. Comparison of N99 and JM2.300

Discussion

N99 cells (CI+) showed higher fluorescence intensity than JM2.300 cells (CI-). From this experiment, we assume that our rm/lac hybrid promoter is actually activated by CI.
We are now confirming that our rm/lac hybrid promoter is not only activated by CI but also repressed by LacI.

For more information, see [http://2013.igem.org/Team:Tokyo_Tech/Experiment/RM-lac_Hybrid_Promoter_Assay our work in Tokyo_Tech 2013 wiki].

Applications of BBa_K1139150

User Reviews

UNIQ0cbc51ce9e4ff8b6-partinfo-00000001-QINU UNIQ0cbc51ce9e4ff8b6-partinfo-00000002-QINU

@@ Line 21: / Line 21: @@
 Fig. 2 is the extracted data which shows the comparison of N99 (IPTG+, with constitutive CI) and JM2.300 (IPTG+, without CI).<br>
-[[Image:titech2013_parts_K1139150_Fig2.jpg|thumb|left|300px|<b>Fig. 1.</b> Fluorescence intensity detected by flow cytometer]]
+[[Image:titech2013_parts_K1139150_Fig2.jpg|thumb|left|310px|<b>Fig. 1.</b> Fluorescence intensity detected by flow cytometer]]
-[[Image:titech2013_parts_K1139150_Fig3.jpg|thumb|none|300px|<b>Fig. 2.</b> Comparison of N99 and JM2.300]]
+[[Image:titech2013_parts_K1139150_Fig3.jpg|thumb|none|240px|<b>Fig. 2.</b> Comparison of N99 and JM2.300]]
 ===Discussion===