Team:Cornell/Notebook/Salicylate reporter/July15

From 2012.igem.org

(Difference between revisions)
(July 16th)
(July 15th-21st)
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=July 15th-21st=
=July 15th-21st=
==July 15th==
==July 15th==
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Dylan noticed that we had colonies on the plates re-streaked from the original plates of conjugated S. oneidensis. He started overnight cultures of S20 (strain list) in kanamycin so that we can sequence and make glycerol stocks. He also started overnight cultures of w.t. S. oneidensis to innoculate into new reactors in Riley Robb, which will be used as positive controls.
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Dylan noticed that we had colonies on the plates re-streaked from the original plates of conjugated S. oneidensis. He started overnight cultures of S20 ([http://2012.igem.org/wiki/index.php?title=Team:Cornell/Notebook/StrainList&oldid=28141| strain list]) in kanamycin so that we can sequence and make glycerol stocks. He also started overnight cultures of w.t. S. oneidensis to innoculate into new reactors in Riley Robb, which will be used as positive controls.
==July 16th==
==July 16th==
The overnight cultures of S. oneidensis need more time to grow - overnight cultures of these leisurely bacteria should be started at noon the previous day, instead of in the evening as with speedy E. coli.
The overnight cultures of S. oneidensis need more time to grow - overnight cultures of these leisurely bacteria should be started at noon the previous day, instead of in the evening as with speedy E. coli.
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Dylan ran another Phusion PCR of the entire nah operon, since we got a lot of mispriming the first time we ran it. The thought was that we had run the previous PCR at the optimal temperature for Vent but used Phusion, so Dylan set up the new PCR correctly, with a single annealing temperature of 66degC and a lengthened final extension time of 15 minutes to account for the size of the nah operon (~10kb). Caleb ran a gel of the PCR and visualized a single band ~9.5kb. PCR of the nah operon out of p20, P. putida ([http://2012.igem.org/wiki/index.php?title=Team:Cornell/Notebook/StrainList&oldid=28141| strain list]) was successful!
[[Image:2012_07_16_nahOperonYES.jpg|thumb|Success!]]
[[Image:2012_07_16_nahOperonYES.jpg|thumb|Success!]]
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==July 17th==

Revision as of 14:36, 17 July 2012

Home Team Project Parts Modeling Notebook Protocols Safety Attributions

Contents

The salicylate reporter is analogous to the arsenic reporter, but with a salicylate-sensitive promoter. That is, in the presence of salicylate it will produce MtrB, completing the Mtr pathway and allowing Shewanella to produce current in our biosensor. In combination with the nah operon, this reporter will be able to detect naphthalene levels. Back to salicylate reporter week view

July 15th-21st

July 15th

Dylan noticed that we had colonies on the plates re-streaked from the original plates of conjugated S. oneidensis. He started overnight cultures of S20 (strain list) in kanamycin so that we can sequence and make glycerol stocks. He also started overnight cultures of w.t. S. oneidensis to innoculate into new reactors in Riley Robb, which will be used as positive controls.

July 16th

The overnight cultures of S. oneidensis need more time to grow - overnight cultures of these leisurely bacteria should be started at noon the previous day, instead of in the evening as with speedy E. coli. Dylan ran another Phusion PCR of the entire nah operon, since we got a lot of mispriming the first time we ran it. The thought was that we had run the previous PCR at the optimal temperature for Vent but used Phusion, so Dylan set up the new PCR correctly, with a single annealing temperature of 66degC and a lengthened final extension time of 15 minutes to account for the size of the nah operon (~10kb). Caleb ran a gel of the PCR and visualized a single band ~9.5kb. PCR of the nah operon out of p20, P. putida (strain list) was successful!

Success!

==July 17th==