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Team:UEA-JIC Norwich/Weekten
From 2011.igem.org
Monday 15th August
The Moss party consisting of Alistair, Abbie and Ben Hardy commence the day by transferring grown protoplasts to new plates; protoplasts containing the Ble gene were subjected to Ble selection plates. The Moss party also performed a PCR on their Nos terminator, and then ran a gel to check it was at the correct size; later to be followed up by gel extraction. The Gel image shows something is there, just perhaps not what we were hoping for though.
Tuesday 16th August
Mario and Mark today tested the G-luc plasmid in transformed E.coli grown in LB cultures. The cultures were grown to the ideal phase and then the cells were sonicated to allow the substrate to be be subjected to the enzyme for an optimum reaction rate. The reaction was observed in the Night-owl machine and showed successful bioluminescene concluding the plasmid worked as suggested.
Wednesday 17th August
Alistair, Abbie and Ben Hardy today mini-prepped the green and yellow fluorescent protein Biobricks from transformed E.coli ready for some promoter testing and to see if they are viable to use in Moss.
Thursday 18th August
Kim made some algae cultures, preparing for more future algae transformations. Ben Jevans and Gurdeep conducted a mini-prep on transformed cells supposedly attaining the Ble gene. They also mini-prepped G-luc transformed E.coli cells grown from two different cultures, one containing the antibiotic ampicillin and one without ampicillin. Results of the mini-prep’s were ran on a gel and then nanodropped; the final outcome of these look fairly promising.
Friday 19th August
The team eagerly start the day with Kim streak plating algae colonies on TAP plates. These colonies were taken from originally transformed plates with algae containing the circular G-luc plasmid and the linearized G-luc plasmid. Ben Jevans throws himself into another PCR reaction hoping to amplify the G-luc and arginine bio-synthesis genes. The final product of the PCR was then ran on a gel; results today don’t look so desirable. Abbie, Alistair and Ben Hardy transformed the double terminator Biobrick and the kanamycin resistance cassette into E.coli, then left it in incubation to grow and use another day. The trio followed this up by a restriction digest on the CaMV promoter and chloramphenicol plasmid PCR products which was then ran on another gel. Results looked okay, however this was then followed by a ligation which was ran on another gel and this time the ladder was the only DNA on the gel to make an appearance. Today half the team take a leave for a short holiday; they will reappear in a couple of weeks whilst the other half stick it out until there return.
Saturday 20th August
Alistair, Abbie and Ben Hardy show some true commitment by coming in on the weekend to do some mini-prep work on the transformed cells containing the desired ribosome binding site. This was followed by a nanodrop to re-assure a sufficient concentration of DNA was available. The trio then made a liquid culture of the supposedly dubious ligation product of the following day and the kanamycin resistance cassette Biobrick.
