Arthur Yu Notebook

From 2007.igem.org

Revision as of 21:59, 12 June 2007

My Construction Files
My Sequencing Files

6/12 a bag full of grapes

• YAY WE ALL GOT OUR OWN SET OF PIPETTE PEOPLE
• PCR of yfbE...

1. last night's thing, left in the freezer overnight. >FAIL<
2. Did a new PCR -- looks good -- cells xform'd, plate is incubating.

• neuS new xformation looks good. Three colonies now incubating.
• wbbL (1) and HPI/katG (4)

1. miniprepped and digest gel ran:

1. HPI/katG 1,2,3,4 || wbbL || marker
2. 1,2 might be okay.. that faint band is weird. 3 is great! 4 = wtf. wbbL = wtf too (should have two bands)
3. decision to put 1,3,4,wbbL for sequencing.

Ayu 17:59, 12 June 2007 (EDT)

6/11 austin's birthday

• CAKE PARTY - great custard cake
• I put the wbbL (1) and HPI/katG (4) colonies to incubate in LB broth.
• neuS failed; no colonies :(((((((
  • redid ligation and xformation. hopefully there will be good results tmrw!
• made like 20 LB-Agar/Amp plates - looks like our stock will last at least this week
• researched nitric oxide (NO) and E. Coli - looks like soxRS is promising
• also researched RBCs and how they deal with NO
• plopped yfbE into PCR will do stuff with it tmrw

TO DO: enter yfbE into the registry
Ayu 18:24, 11 June 2007 (EDT)

6/8 long day?

• My PCR from last night (HPI/katG) was ROXOR! (left)
  • xformed some DH10B's. w00t w00t
• Today's PCR was wbbL and neuS. ALSO ROXOR LOL (right)
  • xformed DH10B's.
• made oligos for yfbE promoter thingy - will test with GFP and yeah! next week!
• poured lotsa LB/agar+amp plates

6/7 we got benches

• we got benches
• pcr of [http://partsregistry.org/Part:BBa_I716253:Design HPI/katG from Salmonella]

1. well... getting the mutated PCR prod overnight. going to xform tmrw, hope it works!

• programmed pcr on machine upstairs (#6)
• we got computers
• AGAR SUX, for future reference:

1. nuke @ 20:00 min, 50% power.
2. water bath in tap water for 5-10 min
3. thaw the antibiotic right now!!
4. FIRE for disinfecting
5. pour that stuff. set 15 min, then marker it then bag

6/6 waiting for oligos

• Made oligos and constructs with Vai, for getting wbbL and neuS from pJ23006-Bca9106
• We tried the P_tet/RFP triple/double digest to make a composite part.

1. FAIL
2. probably source DNA is bad
3. so much for that activity...

Other stuff: I won speed scrabble. even though I kind of cheated ish (didn't stop when Sam said stop"

6/5 coolbeans

• Finalized oligos to order with Vai
• Learned about LB broth-ing and LB/Agar plating. Thanks, Austin and Sam :)
• Learned about the many composite part-making methods. Props 2 Chris

1. prefix/suffix is weaksauce
2. Use the AlwnI or BsaI or BglI, in conjuction with BglII or BamHI << (Did this today)
3. DBBS
4. 3 antibiotic; MIT endorses, used for BioBrick 1.0. Triple digest = bad
5. 1-2-3 method << 'Our Goal' in a few weeks. should be leet.

• Planned and vicariously did the making of P_tet+RFP brick (see Vai Notebook)

Other Notes: All oligos are being ordered, w00t w00t. Ayu 18:36, 5 June 2007 (EDT)

6/4 Training Finishes, Real Stuff Starts

• Incubated some colonies
• Miniprep'd already-been-incubated colonies (2)
• Double digest of the 2 minipreps + parent plasmid
• Colony PCR'd the incubated E.coli
• Ran gel of the digest + PCR

• >>> PCR product / Miniprep 1 / Parent Plasmid / Miniprep 2 / Ladder >>>
• No bands for PCR or parent. Confused? Other ones look great.

As for me: Wiki acc works now.
Designing oligos and will compare with Vai. Ayu 18:19, 4 June 2007 (EDT)

to do