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Team:Cornell/Week 6
From 2011.igem.org
Week 1 | Week 2 - 5 | Week 6 | Week 7 | Week 8 | Week 9 | Week 10 | Week 11 | Week 12 | Week 13 | Week 14 | Week 15 | Week 16 | Week 17 | Week 18 | Week 19 | Week 20 | Week 21 |
July 10th - July 16th
Sunday, July 10
Lab work done by: James Mathew, Charlie Chung
- Worked on website
- - Preliminary design of banner, safety content, and temporary lab notebook spreadsheet
- Ligation product was transformed in electrocompetent cells
- Culture plated on agar plate treated with ampicillin
- Plasmid containing the Vio operon was transformed and plated on agar treated with kanamycin
- Overnight PCR reaction for RFP
Monday, July 11
Tuesday, July 12
- Finished final design of team logo
Wednesday, July 13
- "Website Team" meeting to continue design and construction of website
- - Designed and constructed final banner design
Thursday, July 14
Lab work done by: James Mathew, Charlie Chung
- Sequencing revealed no GFP in the ligation product
- Will retry the 3-piece ligation of GFP + Annealed Primer Pair A (half of biotin tag) + Annealed Primer Pair B (other half of biotin tag & iGEM suffix)
- PCR to amplify vector backbone containing RFP
- Met Prof. Lucks -- prospective iGEM advisor and coolest man alive
Friday, July 15
- Banner was updated to include Flash animation of moving gears.
Team Meeting
- Ligation didn't work -- retry over the weekend
- - transform and plate today
- - inoculate and colony PCR on Saturday
- - Miniprep and prepare for sequencing on Sunday
- Prepare more culture of backbone plasmid
- - inoculate today
- - Miniprep on Saturday
- Order primers today for other methods of ligation: (1) longer primers (2) two-step PCR with shorter primers
- Order primers for our pathway enzymes: VioA, VioB, VioB
- - Primers should come Wednesday
- - If ligation this weekend fails, we can try these other PCR methods
- Animations and website construction are continuing
Lab work done by: James Mathew, Charlie Chung, Youjin Cho
- Running low on pZE vector backbone, so fired up two cultures
- Miniprep to purify out the plasmid. Followed standard Qiagen Miniprep protocol for microcentrifuge
- Verify if PCR amplification of RFP worked using gel electrophoresis. If yes, then proceed to gel extraction and purification
- - NOTE: Thermocycler did not keep our rxn tube at 4°C
- Electroporation method of transforming our 3-piece ligation product and control into DH5α (competent E. coli = made porous for uptake of outside DNA)
- Plated transformed bacteria onto agar plates treated with ampicillin
Saturday, July 16
Lab work done by: Youjin Cho, Nicholas Kramer
- Growth of numerous colonies suggests the ligation reaction worked
- - Will pick 2 colonies off the plate in the evening and inoculate overnight
- PCR to amplify vector backbone containing RFP
- - Melting temperature of 57°C leads to non-specific binding
- - Ran 2 PCR reactions with melting temperatures 59°C and 61°C
