From 2008.igem.org

Revision as of 15:35, 19 August 2008

Newcastle University

GOLD MEDAL WINNER 2008

Lab Notebook

Meetings The team meets at least once a week on thursday afternoons. Details of each meeting can be found through the calendar links.

Ria and Jess finally started in the labs on Monday the 4th August. So we will start updating the wiki with what's been happening and the progress we are making.

Aim 1: clone the spaRK system from pUC57 into pJWV021 and transform into DH5 alpha competent E. coli

Aim 2: clone the spaRK system from pUC57 into pGFP-rrnB and transform into TOP10 competent E. coli

The 2.2kb fragment (ncl08) contains the spaRK system and a promotor-less GFP gene linked to this. This means that when spaR is activated, its positive regulatory effect on spaK will in turn activate GFP.

Monday 4th August

All 5 of us (Megan, Mark, Nina, Ria and Jess) went into the lab today to decide on a plan of action for the weeks to come.

• We used electrophoresis to check for the presence of plasmids (pGFPrrnB- our plasmid with the GFP gene, and pJWV021- with the mCherry gene). Our gel was inconclusive, so we decided to re-run it the following day to confirm our results.

• In the afternoon we attempted to transform DH5 Alpha competent cells with DNA from the registry (from source plate 1010 well 4A and 1018 7A).

• We followed the extraction method outlined in the green folder and for each DNA spot we plated two petri dishes: one with a larger volume of DNA (4μl) and one with a smaller volume (2μl) to make colony counting easier. We plated the following and incubated at 37˚C overnight:

Plate 1: +ve control (isolated plasmid plus TOP10 cells)

Plate 2: -ve control (TOP10 cells only, no plasmid)

Plate 3: Large 1010 4A

Plate 4: Small 1010 4A

Plate 5: Large 1018 7A

Plate 6: Small 1018 7A

Tuesday 5th August

Today we re-ran the gel electrophoresis of pGFP-rrnB and pJWV021 to check correct plasmid size and sufficient plasmid quantity. Results showed this to be true and the expected plasmid sizes of 8399bp for pGFP-rrnB and 7066bp for pJWV021.

Mark's Lab Journal

Nina's Lab Journal