Team:Queens Canada/Notebook/Week7



Notebook - Week 7

DateProtocolPeopleDNA (if relevant)Quantities and Parameters (if relevant)Notes on Protocol/ Gel Lanes
Jun-12Testing Streak PlatesFaisal, David, VictorUsing RFP  
Jun-13Calculating PCR Dilutions Primers arrived, prepare for PCR  
Jun-14Hotstart PCRVictor, Kevin, Beini, PhillipGFP (G1(1), G1(2)) and Luciferase (L1(1), L1(2) were used as template DNAMastermix (x6 trials)
ddH2O 9 uL
Hotstart 12.5 uL
Front Primer 0.75 uL
Reverse Primer 0.75 uL
Template DNA 2 uL
 Gel GFP and Luciferase 1) 1 KB+ Ladder
2) GC / LC
3) G1(1) / L1(1)
4) G1(2) / L1(2)
5) G2(1)/ L2(1)
6) G2(2) / L2(2)
Jun-15Hotstart PCRFaisal, Andrew, KevinPbrB (Lead Binding Protein), fMt (Metallothopnein binding Arsenite and Arsenic), CFPPCR Mastermix
dd H20 10 uL
Hotstart 12.5 uL
LR 0.75 uL
RP 0.75 uL
Template 1 uL
Total 25 uL
Program 61 degrees celcius 15 cycles
Labbeled P1-P3 (PbrB), F1-F3 (fmT).
 Gel ElectrophoresisDavid, Phillip, Beini, AndrewFor previous PCR 1) 1KB+
2) Blank
3) Blank
4) PC
5) P2
6) P3
7) P4
8) Ladder 1 KB+
9) Control
10) F1
11) F2
12) F3
13) F4
14) P1
15) Ladder 1 KB plus
 Gel ElectrophoresisFaisalGel for CFP and GFP from ealier in the day 1) 1 KB+ Ladder
2) GFP C
3) GFP 1
4) GFP 2
5) GFP 3
6) GFP 4
7) 1 KB+
8) CC
9) C1
10) C2
11) C3
12) C4

Monday June 11

Today we split into various groups and continued research on our respective topics, such as non-BioBrick parts (specifically, potential synthesis pathways for today) and potential fusion standards. In the afternoon, we continued researching the same topics we did in the morning, while Kevin and Phillip had an interview with the local newspaper, Kingston This Week. Phillip and Kevin also updated the team website, with this year's content.

Tuesday June 12

Today we again split into various groups and researched our different topics, with a particular focus on applications of our project. We also spent a lot of time doing sponsorship, and we got our first sponsorship deal! *Insert Fanfare here*. From the Queen's Chemical Engineering department. Phillip also spent a lot of time working on the wiki, finally getting a clear picture of the navigation menus on the site. Kevin and Faisal had a meeting with one of our advisors, Dr. Campbell, to discuss modelling the chimeric flagellin. We also uploaded some of the pictures that we had from yesterday's SynthetiQ work (thanks to Beini for the captions), and Kevin had a skype conversation with the inspiration behind SynthetiQ: John Bohannon!

Wednesday June 13

In preparation for the arrival of our primers, which are due to arrive tomorrow, we spent most of our day researching PCR techniques and protocols. In particular, we researched PCR troubleshooting, in case anything goes wrong, which hopefully it will not. During lunch, we went to a workshop in which we received a list of things to make work more fun (if that's even possible), and Andrew hopes to do them all before the summer ends. Coincidentally, one of the items on the list was "Eat perfectly ripe grapes," which we did yesterday, thanks to Mitangi. In the afternoon, we met with our faculty advisors in order to both discuss our progress as well as get more information on the financial side of our project. In the lab today, we worked on some bacteria art to get some cool pictures for when Kingston This Week will come and take pictures on Friday.

Thursday June 14

Today, we spent most of the day doing research on making our own restriction enzymes and working on sponsorship. We also spent time in the lab today, and did two things: analyze the results of yesterday's bacteria art (which turned out well), and perform Hotstart Ready PCR for GFP and luciferase. After PCR finished, we performed gel electrophoresis on both parts, and based on the results, they both worked.

Friday June 15

In the morning, a photographer from Kingston This Week came to take photos of us for the article about us. After that, we split into two groups: one in the lab performing PCR for fmT and PbrR, and the other spending more time researching the creation of our own restriction enzymes and finishing off the discussion about which BioBrick fusion standard to use. At the beginning of the afternoon, we finally heard back from OSLI.....and we got it! Our second sponsor! After that, we split into groups in the lab. Some worked on gel electrophoresis for GFP and luciferase, others worked on PCRing GFP with linkers and ECFP, and William worked on determining the error on our pipettes. It was a real party in there with all of us in the lab. After PCR was finished, Faisal and Kevin performed gel electrophoresis on the remaining two BioBrick parts. In the end, the results showed that PCR worked for fmT and one of the GFP with linker gels showed up. The PCR for PbrR did not work as nothing was shown. The PCR for ECFP showed a lot of gels lower down the ladder. It is possible that the plasmid was not taken up for the lead-binding proteins and the ECFP proteins; these two colonies might have to be regrown.