"
Team:Cornell/Notebook/Salicylate reporter
From 2012.igem.org
| Home | Team | Project | Parts | Modeling | Notebook | Protocols | Safety | Attributions |
|---|
|
Write description
June
Week
June 29th, Friday
- Vent PCR at 11:00 (DPW)
- Amplifying both previous Phusion PCR band and original p21 template
- Dylan's magic triple anneal method (55/60/63)
- Gel purified PCR product from Phusion template (~1:20 pm) (DPW)
- Quantified product at 22.4 ng/uL
- Set up digestion of p21 PCR product with EcoRI-HF and AscI (~9:00 pm).
- 22 ng/uL --> 45.5 uL sample for 1 ug digest
- Buffer 4
- Ran digestion on gel. (~11:00 pm)
- Sliced out relevant band on gel, stored overnight at -20.
- Miniprepped overnight cultures of PL14-PL20 (~1:00 pm, STC)
- Using C1015 rotor, 6666 x g, the Corning culture tubes only fit in the centrifuge with the lids off
- Made LB, 3x 60 mL in 100 mL bottles (~ 3:00pm, SS)
- Made LB Agar, 4 x 250 mL LB Agar in 500 mL bottles (~3:00, CS)
- Autoclaved LB, LB Agar, and milliQ (~3:30 pm, SS)
- Made LB plates with Kan (~6:30 pm, CMR)
- CUGEM movie outing at 8:00 pm.
- Phusion PCR at 10:00 pm (DPW)
- Dylan's magic triple anneal method (57/65/70, 35 cycles total)
- Amplifying nah operon from Gibson 1
- Appending BioBrick cutsites for ligation into pSB3C5
June 30th, Saturday
- Took PCR out of thermal cycler at 9:00 am (DPW)
- Set up gel using NEB 100bp and 2-log ladders (10:00am)
- Continued gel extraction of p21 PCR digest from previous day (SS)
- Set up ligation of p21 PCR digest extraction and dephosphorylated pBBRBB+mtrB (11:11am, SS)
