Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2014/05/22: Difference between revisions

From OpenWetWare
Jump to navigationJump to search
Line 20: Line 20:
| EsaI||0.046||0.026||1.776||45.692
| EsaI||0.046||0.026||1.776||45.692
|}
|}
<br><br>
picked all the colonies on the LuxR, RhlR, EsaR golden gate receivers with the 10:1 ratios and streaked them on a new plate. dotted 8ul of sender cells in the center of the plate. put at 37°C at 11am.
<br><br>
<br><br>
Going to run golden gate anyway. Backbone plate should have the same number of PCR template transformants as any of the ligation plates.  
Going to run golden gate anyway. Backbone plate should have the same number of PCR template transformants as any of the ligation plates.  
Line 41: Line 43:
| EsaI||0.77||1.9||0.5||1.25||2||13.58
| EsaI||0.77||1.9||0.5||1.25||2||13.58
|}
|}
<br><br>
Set up PCR of LuxI with P131 and P132 AT 50°C ext time 1 min because it failed the other day with an annealing temp of 50. 


<!-- ##### DO NOT edit below this line unless you know what you are doing. ##### -->
<!-- ##### DO NOT edit below this line unless you know what you are doing. ##### -->

Revision as of 15:44, 22 May 2014

Project name <html><img src="/images/9/94/Report.png" border="0" /></html> Main project page
<html><img src="/images/c/c3/Resultset_previous.png" border="0" /></html>Previous entry<html>&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;&nbsp;</html>Next entry<html><img src="/images/5/5c/Resultset_next.png" border="0" /></html>

mm/dd/yyyy

PCR clean up of EsaI PCR and mCh Sender Intermediate for golden gate. Forgot to do DpnI digest of mCh (don't need to for EsaI because the PCR template is kan resistance).

Sample 260 280 260/280 ng/µL
mCh Sender int gg 0.055 0.031 1.774 55.337
EsaI 0.046 0.026 1.776 45.692



picked all the colonies on the LuxR, RhlR, EsaR golden gate receivers with the 10:1 ratios and streaked them on a new plate. dotted 8ul of sender cells in the center of the plate. put at 37°C at 11am.

Going to run golden gate anyway. Backbone plate should have the same number of PCR template transformants as any of the ligation plates.
200fmol of insert, 20fmol backbone

Reaction Rec Bb LuxI BsmBI T4 Ligase T4 Ligase buffer H2O
Bb ctrl 0.77 0 0.5 1.25 2 15.48
RhlI 0.77 3.2 0.5 1.25 2 12.28
LasI 0.77 2.8 0.5 1.25 2 12.68
EsaI 0.77 1.9 0.5 1.25 2 13.58



Set up PCR of LuxI with P131 and P132 AT 50°C ext time 1 min because it failed the other day with an annealing temp of 50.



Retrieved from "https://openwetware.org/mediawiki/index.php?title=Haynes_Lab:Notebook/Characterizing_AHL_quorum_sensing_homologs/2014/05/22&oldid=794607"