Team:Cornell/Notebook/Nah operon


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Project Description

The purpose of this subproject is to make the nah operon biobrick compatible for the use of future iGEM teams. The nah operon is an operon that converts napthalene into salicylate. In our system, this will allow napthalene to be detected by the salicylate reporter. There are three standard cutsites internal to the operon which we are removing - the biobrick compatible piece could be used by future teams in bioremediation projects.

June 13th, Wednesday

Transformed pS

Set up PCR for four fragments of the nah operon as well as the pBMT-1 backbone, using primers designed to mutate cut-sites concurrently with Gibson Assembly. Also set up a PCR for the entire nah operon; if Gibson Assembly fails, but we are able to PCR the entire 10kb nah operon using Phusion polymerase, an alternate method of mutation using Phusion will be pursued.

June 14th, Thursday

June 18th, Monday

We will be finishing the Gibson assembly of the nah operon in Olin tomorrow. For now, plan on meeting at 301 at 9am. It's possible that we'll have to push it back for Taylor's convenience, and I'll let you know if that happens. The process shouldn't take incredibly long, since the Gibson method just requires a one hour incubation... and then we just need to transform into cells.

Tomorrow, Claire et al. will also be testing out the electroporator in Weill so that we can transition to the BME lab space. This will be happening after the Gibson stuff in finished in Olin. So... you can venture to Weill if you are excited about electroporation.

June 19th, Tuesday

We ran the Gibson assembly and transformed it into cells, which are incubating right now. We're also running PCR to amplify the Gibson products. Taylor's setting up a digestion of the PCR products tomorrow morning, and Dylan and I will be in Olin tomorrow around 2 to run the digested products on a gel to see if Gibson worked. We'll also check to see if we got colonies from the transformation then.

June 27th, Wednesday

  • Ran digest of gibson-assembled nah operon on gel

===June 28th, Thursday===