Haynes Lab:Notebook/Characterizing AHL quorum sensing homologs/2013/05/30: Difference between revisions

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Supernatant transfer:
Supernatant transfer:
Spun down 4ml of culture in 2-2ml tubes for each sender. Poured supernatant into round bottom tubes. Dropped some supernatant onto pH strip. Added 1ml of receiver cells to a 2ml centrifuge tube. Added 20ul of Amp and shook. Added 100ul of Receivers + (~4ul) Amp to each 4ml volume of Sender cells in round bottom tubes. Added 1ml of LB+Amp to add nutrients and lower the pH a little. Same Sender-Receiver pairs as above. Started shaking ~10:30.
Spun down 4ml of culture in 2-2ml tubes for each sender. Poured supernatant into round bottom tubes. Dropped some supernatant onto pH strip. Added 1ml of receiver cells to a 2ml centrifuge tube. Added 20ul of Amp and shook. Added 100ul of Receivers + (~4ul) Amp to each 4ml volume of Sender cells in round bottom tubes. Added 1ml of LB+Amp to add nutrients and lower the pH a little. Same Sender-Receiver pairs as above. Started shaking ~10:30AM.
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Determine pH using test strips:
Determine pH using test strips:

Revision as of 10:31, 30 May 2013

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Entry title

9AM

BL21 sender cultures grew in 6mls and 2mls. RhlR-GFP still didn't grow.
Shook LuxR, TraI, and TraR that had been in the fridge for a few minutes.

Co-cultures: Add 100ul of Sender, 100ul of Receiver to 3ml LB+Amp (except for TraI culture, which will not have antibiotic). Started shaking at 10AM.

Sender Receiver
LuxI LuxR
LasI LuxR
RhlI LuxR
TraI LuxR
TraR ctrl LuxR



Supernatant transfer: Spun down 4ml of culture in 2-2ml tubes for each sender. Poured supernatant into round bottom tubes. Dropped some supernatant onto pH strip. Added 1ml of receiver cells to a 2ml centrifuge tube. Added 20ul of Amp and shook. Added 100ul of Receivers + (~4ul) Amp to each 4ml volume of Sender cells in round bottom tubes. Added 1ml of LB+Amp to add nutrients and lower the pH a little. Same Sender-Receiver pairs as above. Started shaking ~10:30AM.
Determine pH using test strips:

Sender pH
Plain LB 6-7
LuxI 7-8
LasI 7-8
RhlI 7-8
TraI 7-8
TraR ctrl 7-8





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