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Team:The University of Alberta/16 June 2008
From 2008.igem.org
Troubleshooting Day
We have been getting alot of bad results (or no results at all) in the lab lately - our gel purifications have had very low yields, the Westerns have no been working, our transformants wont grow, etc. We have decided that the next few days will be dedicated to troubleshooting so we can figure out what's going on, or what isn't going on. Troubleshooting tasks have been divided up thusly:
Jason
- Digest reporter genes from pUC57
- gel purify
- sequence
- Digest biobricks from J61003
- gel purify
- sequence
This will let us know two things: if the BioBricks we currently have in J61003 are actually the biobricks we want and if the original genes we were given in pUC57 are actually the reporter genes they are purproted to be. This could help explain why our Purple Russian colonies aren't purple, and why our PCR results are always so sketchy.
- Retransform biobrick ligations into XL-10 Gold competent cells
Our transformants havent been growing for some reason so we are goign to try and transform them yet again. We will also try tansforming them into the commercially prepared cells to compare
Tom and Winnie
- Set up O/Ns of butanol stuff for protein expression
- Use Ni-NTA columns to purify
- Run on SDS-PAGE gel to confirm
- Sequence, PCR, digest, confirm butanol biobricks
This should help us understand what's going on with our butanol stuff (why they dont seem to be growing in I0500, etc)
Chris
- Redo Westerns
Our westerns have been totally blank and we dont know why. Redoing them, in conjuction with Tom and Winnie's stuff above, may shed some light into why they have not been working.
Saima
- Make 3 gels - one from each of the 3 sources of agarose in the lab - then run a standard and gel purify. This will let us know if the agarose we have been using is the cause of the low yields from gel purifications.
