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Latest revision as of 17:23, 18 September 2011

July 17th - July 23rd

Sunday, July 17

Lab work done by: Alyssa Henning, Bill Jo

• Miniprepped 1 sample of GFP + AviTag (the second sample got messed up)
• Ran a gel on the RFP that was amplified via PCR on Saturday
• Gel purification and extraction
• Sean will inoculate 6 tubes of GFP-containing bacteria -- 1 tube for us, 5 tubes for CURIE (Cornell iGEM's outreach program)

Monday, July 18

Lab work done by: Charlie Chung, Youjin Cho

• Gel purified 2 RFP PCR samples from Friday
• Miniprep 2 samples of GFP + AviTag

Tuesday, July 19

Lab work done by: James Mathew

• Submitted GFP + AviTag for synthesis
• Submitted pZE12 backbone for subcloning of light sensor

Wednesday, July 20

Lab work done by: James Mathew

• Set up ligation reaction of pZE-12 backbone with the primer dimer insert

1. Digestion of Backbone

- 26µl Backbone plasmid = 2µg DNA

- 5µl NEBuffer 4 (optimal for SphI-HF and ClaI-HF)

- 1µl SphI & 1µl ClaI

- 16.5µl H2O

• left in water bath for one hour

- 1µl CIAP added to digestion reaction

• left in water bath for 30 minutes

2. Ran Digestion Product through gel for 30 minutes at 120V

Lane 1: 2µl 1kb Ladder + 2µl 6x loading dye + 8µl H2O

Lane 2: 25µl digestion reaction + 5µl 6x loading dye

Lane 3: 25µl digestion reaction + 5µl 6x loading dye

3. Gel Purification using Qiagen Kit

4. NanoDrop of Samples

5. Ligation Reaction

• Ligation done overnight for transformation on 7/21/11

Thursday, July 21

Lab work done by: James Mathew

• Miniprepped Vio operon (Cambridge 2009)

Lab work done by: Youjin Cho, Charlie Chung, Alyssa Henning

• Reconstituted VioA, VioB, and VioE forward and reverse primers
• Set up PCR for VioA, VioB, and VioE
• Aborted transformation of pZE12 plasmid + AviTag primer dimer because reverse primers for AviTag were incorrect
• Redesigned reverse primers for AviTag to order on Friday

Friday, July 22

Lab work done by: James Mathew, Claire Paduano

• PCR gel purification of VioA, VioB, and VioE

Saturday, July 23

Lab work done by: James Mathew, Claire Paduano

• Digestion of VioA, VioB, VioE, GFP, and RFP in preparation for ligation with corrected backbone and AviTag primer dimer