Alberta/Calender/September
From 2007.igem.org
September
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September 1
Lab Notes
1. 24 Minipreps for Diezel Blaze in B0034 using Fermentas Kit; stored in -20 freezer
2. Lots of colonies from the transformation last night; no colonies on negation control (good) and lots of colonies on positive control (good) and no satellite colonies (also good); no colonies were picked for overnights
3. Run the Buddy+Benny in B00 and Betty+Enny in B00 Pst/Xba gel from last night at 50V for another 30 minutes for better resolution; all the digest worked so I cut out the appropriate bands and stored the gel in -20 freezer for purification later
-CZ
September 2
September 3
September 4
Schedule:
JP,ML,JG 7:00pm
For the record:
Buddy = BuOH DH 1235 bp
Betty = Bb-bhBu-coa dh 923 bp
Benny = Butyryl coa dh 1184 bp
enny = Enoyl coa Hydratase 860 bp
Diezel Blaze = buAld-Dh 2639 bp
Notes:
Moving was done this mid-day.
New location M349
JG has key & access
- MC, ED, JG
Evening crew - please do a once over of G308 to double check we have not left anything and to clean up anything extra/ things we should get out of the way. thnx - MC
Night Crew
-JP, JG
Set up new lab
Things we need - Water Bath, Scale, disposable 13ml overnight tubes, Tip waste
lab work
Gel purification ran on Betty,enny and boo as well as buddy and benny and is in the -20 in the new lab
To Do list for tomorrow
- Restriction on DB needs to be done - If we want to put it in front of BE or BB then Spe/Pst - If we want to put it behind BE or BB then cut with Xba/Pst - maybe do both (use 4microL DNA for digest)
-complete restriction on BE and BB for SPE and PST (then we can complete ligations BE-BB and BB-BE or we can ligate D.B into BE or BB to make BE-DB or DB-BE or BB-DB or DB-BB- Then we can ligate BE-BB-DB or DB-BE-BB or BE-BB-DB or BB-DB-BE or BB-BE-DB and we are kinda finished!
- If I0500 comes in from calgary it needs to be transformed.
- Transform R0080 which is a back up promoter just in case?
-Note we also have to do sequenceing on BB and BE and DB
Shout out to calgary for saving our necks Shout out to the move in crew to the top
September 5
Schedule:
NG,ED,ML 7:00pm
AM Crew Notes:
finished up cleaning up post-move in
got a 'water bath' - JG calibrated it
prepared BB in Boo & BE in Boo for sequencing - this will be ready for pick up tomorrow
Restricted:
- DB in BOO with PST/XBA
- DB in BOO with PST/SPE
- BB in BOO with PST/SPE
- BE in BOO with PST/SPE
NB: there is a section in the masterbook "figuring stuff out" just a few questions I have from being away so long - don't worry about it we should do a wet lab recap to bring everyone on the same page at our meeting Thrusday - MC
<3 JG & MC
Transformed I0500 from Calgary. Transformed R0080. Ran digestions on gel. Ligated digested DB with BB and BE. (all in Boo)
-ED, ML
September 6
Schedule:
NK ED AL 5:00pm (Unless your crazy enough to show up at 5AM)
To Do:
Ligations
Goodies will be served during the meeting (7:00pm @ CAB 373), so be sure to show up!
September 7
Schedule:
AM Crew: MC, JG meeting at 800hrs
Lab Notes:
No growth observed for R0080 overnights; just scrapping it because we have I0500
No growth on transformations of BB+DB and BE+DB ligations therefore . . . religated - see masterbook for more details
O/N with AMP for I0500
Transform religations in to XL10 Gold
-MC, JG, ML
NB: JG has key access
To Do:
Mini preps on I0500 overnights
September 8
Schedule:
Super Awesome Marathon of Productiveness
8-11:
JG,NG
Miniprep I0500
DB BE and DB-BB transformations did not work.
Re-ligated DB into BB boo and DB into BE boo
11-2:
AL,ML
Re-transform BE+DB and BB+DB
2-5:
VH
Gel extraction of I0500, but we missed the step "restricting with Ecori+speI"
5-8:
JP,NK
Restrction on I0500 mini's with Ecori and SPeI
September 9
Schedule:
And Yet another super productive Sunday
8-11: NG and CZ
NG - Nobody Called me about the key last night (and I completely forgot to swing by: was with parents). So I will wait by the lab either , until the next group shows up or someone with a key. Also bioSci is locked but there is an open door by the physcology wing. I have my cell on 902-8032.
NG - Update: 9:30am 'broke' into lab :D. Then found key...
11-2: VH, CZ
Nick ran the gel of I0500 restricited by Eco/Spe, but nothing showed up as expected (1.5kb).
Digested I0500 and J61003 with Eco/Pst as instructed.
2-5: MC, JG
ran gel of restricted I0500 & J61003; unfortunately there is no DNA in the I0500 sample either
gel extracted J61003
Made LB - to be divided into bottles & autoclaved in G308
O/N of DB in BB & DB in BE samples left in 37degree room
5-8: ML, ED autoclaved little bottles of LB. NOTE: does not need to be autoclaved before you pour it into little bottles and then re-autoclave. Just pour into bottles and autoclave once digested all in Boos except for DB with Pst/xba
September 10
Schedule: And Scheduling just got hard core
I have sheduled what we have taken down in the meeting some people appear many times on this schedule working alone just because of the time they were availible. If you are comfortable working alone and taking on lots of shifts im sure the team would be greatful but I'm also sure we would also understand if you didn't want to work alone in the lab. I have only scheduled this way because we need man hours so i tried to fill up all the spots as best i could. -JG
MC,JG for the AM
put away bottled LB
Ran gel of last night's restricted samples "in boo"; did not really turn out
ran gel of uncut & single restricted I0500; uncut I0500 showed 2 bands and restricted showed nothing
Mini prepped Ligations of DB in BB and DB in BE
to do:
- run gel of double digested I0500
- do single & double restriction on Mini prep of Ligations
- run gel of uncut, single & double restrictions of mini prepped ligations
ML,AL for the MID (AM crew: What time will you guys be at the lab till? I won't be there until 12:30. Might need to arrange something in order to pick up keys. - AL)
Double digest of BB-DB/BE-DB XBA/PST
NG (2:30-4) for the PM (celine is in calgary if you dont feel comfortable in the lab by yourself I can show up - JG)
Jason if you have time to show up that would be sweet, but if you don't I understand. -NG
Ran gel of the restrictions made in MID, Sept 10 by ML and AL.
JP,ED for the Night
September 11
Schedule: No one else really signed up for tuesdays so anyone who has time to drop in should
AM- NK Restrictions of BB+DB with XBA/PST and BE+DB with PST, SPE
Mid - AL (12:30 - 2 ish), NG (random)
Ran gel on restrictions made by NK in AM of Sept 11
Night- ED JP
Gel extractions
Ligations of DB+BB + BE and DB+BE + BB
mini relocation 1 bench back, do not use the first 3 benches of the lab or the blackboard from now on!
September 12
Schedule:
NG Hi guys I have a meeting 3:30 that cannot be reseached I moved myself to come in Tuesday instead (when nobody was in).
AM -
Mid- ML, AL
Important: Please evacuate lab before 2:00pm as there is a lab class starting at this time till 5pm! -AL
PM - VH
lab class 2-5
VH: please make note of the above announcement regarding lab class at 2:00pm
Night - CZ, ED
Transformed BEDBB and BBDB into XL 10 gold
September 13
Schedule:
AM-Nik
Digested BEBOO and BBBOO with PST and XBA
PM - VH, JG
Ran gel of restrictions made in the previous shift.
Meeting 7:00
After lab shift, JG, JP
Overnights with AMP of BBDB + BEDB
Gel extractions from gel made in the prevous shift
bands 1,2 (BE) correct while band 3 is too large to be BB
Sequence reactions can now be started using primers 1-5 that have been diluted to 5pmol/microlitre in H20 not TE
September 14
Schedule:
AM - MC (around 900/930hrs)
- Mini prep from O/N of BBDBE & BEDBB
- Glycerol stocks of BBDBE & BEDBB, and restreaked quartered plates
- Gel extraction of BE & BB Xba/Pst
- made gel
Mid - AL, JG
Restrictions on BBDBE and BEDBB with xba/pst and spe/pst
PM - VH, CZ
Ran gel on restrictions on BBDBE and BEDBB with xba/pst and spe/pst
Gel extracions
night - JP
Sequencing reactions
BBDBBE 6 reactions
BEDBBB 6 reactions
BB 3 reactions
BE 3 reactions
General note: Each primer sequence will be the end of the indicated gene to show us wether the joining/ligations worked correctly (VF is the promoter/gene/juction)
To do: Submit to MBSU
September 15
Schedule:
And yet another crazy weekend...
8-11 - ED,MC
General Note:We have cloned all 5 genes in series, in to differnt orders. Waiting for I0500 which couls be completed on monday. To make sure we have cloned correctly will also need to wait for the squencing results. This weekdn we will clone the 5 gense in 2 differnt orders. The digest have already been done
Gel extraction of gel bands of XBA,PST of BBDBBE and BEDBBB. THese can be used to cone into I0500 J61003
Ligations of DBBB into BE and DBBE into DBBEBB
Leave at 20 degrees celsius for 10 hours
5-8 - NK
Cant find the competent cells.
Competent cells are in -80 freezer 2nd shelf from the bottom-ML, NG
September 16
Schedule: Continued...
8-11 ED, JG
Transform ligations of DBBEBB and DBBBBE into competent cells.
Plated on AMP with whole ligations
General Info: When DBBEBB and DBBBBE are complete run sequence reaction
Follow "flouresence sequence reaction" protocol
11-2 VH, CZ
2-5 MC, JP
5-8 - ML, NG
No growth on DBBBBE or DBBBBE at 1700
Retransformed into competent cells DBBEBB and DBBBBE
Plated 2 of each and lover overnight at 37 degrees
September 17
AM - JG (800 hrs - 930hrs),MC (@ 800hrs)
- delivered MSBU sequencing samples
- O/days of Ligations from yesterday
NB: AMP plates are marked with a red streak down the SIDE of the plate. if it does not have this - then it is NOT AMP
ML (~10:30-1:00)>
September 18
NK - 8 AM
VH - (@1pm)
AL - 12-2pm
September 19
ML - ~10:30 - 1:00
AL - ~1:00 - 2:00
Last Wednesday there was a lab class in our lab from 2-5, is this going to be a regular occurence? -VH
YES - MC
VH, see notes on Sept 12. - AL
ED- 6PM
September 20
NK - 8 AM
VH - (@1PM)
JG, MC- (@5PM)
JP- late
September 21
AM - MC, JG (@ 800hrs),
ML (~10:30 - 1:00)
AL - ~12:30 - 2:00
CZ-2:15PM
ED - 4 PM
September 22
ED 9-12
NG 12-3
NK 230-530
JP evening
September 23
ED 9-12
NG 12-3
NK 230-530
September 24
AM - MC @ 8000hrs
September 25
September 26
September 27
September 28
September 29
September 30
