Difference between revisions of "Part:BBa K398326:Experience"

Latest revision as of 00:56, 12 December 2018

This experience page is provided so that any user may enter their experience using this part.
Please enter how you used this part and how it worked out.

Applications of BBa_K398326

iGEM TU Delft 2010

In order to measure the strength (efficiency) of this promoter we built the part K938331 which is pCaiF+ E0240. Check [http://2010.igem.org/Team:TU_Delft#page=Project/sensing/results our results page] in order to know more about our experience with this part!!!

Condition	Exponential phase [GFP molecules/O.D.]	Stationary phase [GFP molecules/O.D.]

LB	1.2516E+07	-8.6245E+05
0.5LB	1.8101E+07	-2.5945E+05
[glc]=10g/L	6.6456E+06	-6.7109E+06
[glc]=5g/L	8.3673E+06	4.2339E+06
[glc]=2g/L	7.2869E+06	3.9755E+07
Diauxic growth (glucose phase)	7.4517E+06	2.1543E+07
Diauxic growth (Laurate phase)	1.1435E+07	6.6428E+06

Also a simple model for this system was developed. [http://2010.igem.org/Team:TU_Delft#page=Modeling/pcaif-model Click here] in order to know more about it!.

Stanford BioE44 Fall 2018

We placed the 5' regulatory sequence upstream of GFP as a positive control that represents the natural lac operon in action. This part will induce expression when CAP binds (low glucose concentrations).

Due to the clear lack of a trend in fluorescence beyond 0.56 mM for all plasmids and the high variability in results, we chose to repeat this experiment with a narrower range of seven glucose concentrations (1.11 mM to 0.035 mM with two-fold dilutions and 0 mM). There is an indication of glucose-repressed fluorescence for lower glucose concentrations, but this negative correlation does not hold for higher values in this range.

"Time-averaged dose-response curves for Plasmid N from initial time point to three hours with sampling every 15 minutes. Glucose concentrations ranged from 0.035 mM to 17.78 mM with each concentration doubling the previous. Mean ± standard error shown for each point."

User Reviews

In order to convert from AU of fluorescence to GFP molecules we used the conversion factor reported for the part BBa_E0040 which is: 79.429 nM^-1 according to the page of E0040. Nevertheless, we believe that this conversion factor it is also dependent on the path length... We couldn't find information about the path length that the people that characterize E0040 used, thus we are just using the conversion factor ASSUMING THAT THERE'S NO PATH LENGTH DEPENDENCY. In case that you prove that indeed there is a path length dependecy, we use a microtiter plate of 300 uL with a total reaction volume of 100 uL. Then, we ask you to correct the values found.

UNIQ5917d9a3749293a7-partinfo-00000000-QINU UNIQ5917d9a3749293a7-partinfo-00000001-QINU

@@ Line 13: / Line 13: @@
 Also a simple model for this system was developed. [http://2010.igem.org/Team:TU_Delft#page=Modeling/pcaif-model Click here] in order to know more about it!.
+*Stanford BioE44 Fall 2018
+We placed the 5' regulatory sequence upstream of GFP as a positive control that represents the natural lac operon in action. This part will induce expression when CAP binds (low glucose concentrations).
+Due to the clear lack of a trend in fluorescence beyond 0.56 mM for all plasmids and the high variability in results, we chose to repeat this experiment with a narrower range of seven glucose concentrations (1.11 mM to 0.035 mM with two-fold dilutions and 0 mM). There is an indication of glucose-repressed fluorescence for lower glucose concentrations, but this negative correlation does not hold for higher values in this range.
+[[File:PlasmidNExperiment2.png|600px|thumb|center|"Time-averaged dose-response curves for Plasmid N from initial time point to three hours with sampling every 15 minutes. Glucose concentrations ranged from 0.035 mM to 17.78 mM with each concentration doubling the previous. Mean ± standard error shown for each point."]]
 ===User Reviews===
+In order to convert from AU of fluorescence to GFP molecules we used the conversion factor reported for the part [https://parts.igem.org/Part:BBa_E0040 BBa_E0040] which is: 79.429 nM^-1 according to the page of E0040. Nevertheless, we believe that this conversion factor it is also dependent on the path length... We couldn't find information about the path length that the people that characterize E0040 used, thus we are just using the conversion factor ASSUMING THAT THERE'S NO PATH LENGTH DEPENDENCY. In case that you prove that indeed there is a path length dependecy, we use a microtiter plate of 300 uL with a total reaction volume of 100 uL. Then, we ask you to correct the values found.
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