Team:The University of Alberta/26 May 2008

NK 130-530

Hey guys, Kelly and I (Anthony) were in on Friday and we managed to excise the bands and do the gel purification. Kelly has put Winnie's digested producs in the -20, I have put the purification products also in the -20 (wrapped together in green tape and labelled). As well, anything that was left out on the bench was put in the -20 because we have no clue what was in those tubes. Kelly and I were having a difficult time following the lab book, so PLEASE be explicitly clear about what has been performed, what is in each of the biobricks, etc... It might seem annoying and redundant but the volunteers have simply no way of following otherwise. A simple summary of each procedure and protocol would be GREATLY appreciated! We didn't have time to do to the sequencing reaction because Kelly has left for Rocky Mountain, and I am not about to start a 2.5 hr reaction at 7:30 PM on a Friday night!

Good luck with things on Monday, and I will see you Tuesday

Anthony

Today In The Lab
Overday's of Purple Russian and Tryp Finish Fundraising Letter Finish Construction of the Binary Vector  Plant 2 more trays of arabidopsis Work with the Butanol Pathway some more

Jason: I have made overday's of Blue Ox as well as Purple Russian and Tryp.  Finished writing the first draft of the Fundraising letter  Ordered primers for all of our bricks as well as VF2 and VR  Chris:
 * Got the sequences from last week (21, 22, 35, and 99). I tried BLASTing them but I didnt get a single significant match for either part (the only result was a poor match to some expression vector). I tried to do a | Blast 2 on each part to the sequences from last year, but the results were not good:
 * 21: The sequence matched well with the sequence from last year. Did a normal BLAST on last years sequence and it worked fine; makes me wonder why I had no results when blasting this year's sequence.
 * 22: There was no significant match to last year's sequence at all aside from a 41bp match at the start of the sequences.
 * 25: I couldnt do a BLast2 on this part because there was no reference sequence from last year. This part (Butanol dehydrogenase) did have a sequence but it was as a part of a larger BioBrick with another part (butyraldehyde dehydrogenase). Trying to Blast2 these sequences resulted in to significant matches.
 * 99: There was no reference sequence from last year for this part.

Part 23 was not sequenced (did we forget?). Should do this before we do anything with it to make sure its what we think it is! EDIT: Turns out that the parts were mislabelled....What we labelled as 22 is actually part 23 (and 23 is likely to be part 22). This means we have the sequence for the real 23....but we have no sequence for part 22. Tom: Winnie: Kelly and Alyssa S.:
 * Summarized our near future butanol plans on page 37 of the masterbook.
 * Digested I0500 with Spe/Pst and R3298 (butanol DH - I725025) with Xba/Pst.
 * The reason for redoing digestion of I725025 is because last week's band did not match our prediction.
 * Ran the colony PCRs and the two digests above on a gel (similar to the gel ran on May 23 since no picture was taken from that gel).
 * Did NOT run 21 however; could not find the PCR colony for those 4 tubes.
 * Gel excision of I0500.
 * Won't be able to transform the ligations as we are to incubate at room temperature for 3 hours.
 * Did ligation and transformation for I725023 and thiolase
 * set up overnight for I0500
 * Help Tom seting up his gel
 * Did mini-preps of Blue Ox, Purple Russian, and Tryp (4 each). Placed mini-preps in "Mini-Preps 1" freezer box.
 * Made glycerol stocks of Blue Ox, Purple Russian, and Tryp; placed in -80 freezer.

Volunteers Scheduled for Today
5-8 AS 5-8 KR 5-8 SG

Volunteers
<li>If you could would please make Glycerol stocks as well as a mini prep on those Overdays that Jason has done</ul>

A Note To Everyone
There are boxes on the left hand side of the -20 freezer in the lab for our ligations, digestions, mini-preps and random other things. They are labelled "iGEM Digestions 1", etc. If you have anything to store in the freezer, please place items in the appropriate boxes. This makes finding what we want in the freezer a whole lot easier than sifting through racks upon racks of tubes. Don't forget to write down in the lab book (or online) which box you put everything in and where in each box you placed it (we've tried to set up a grid-like numbering system for each box [A1, A2, etc for each space]).