Objective
- Expression of MBP in ER2566
- Redo BSA PCR from pCMV-SPORT6+BSA with correct 3' primers
- Transform QuikChange product from 8/19
Bench work
- Added 500μL 100mg/mL ampicillin to 4 x 500mL 2xYT media in 2800mL Fernbach flask, then added 2.5mL of 40% glucose to each flask as well
- Innoculate two flasks each with 10mL of pMAL-pIII and pMXB10 in ER2566 50mL growths
- Grew 2 hours, then checked OD @600nm (looking for 0.600):
- pMAL-pIII #1 OD = 0.677
- pMAL-pIII #2 OD = 0.646
- pMXB10 #1 OD = 0.622
- pMXB10 #2 OD = 0.639
- Induced expression with 500μL 1M IPTG, grew O/N @ 37°C
- Followed standard Transformation protocol with pMXB10 QuikChange product from Friday
- 3' SapI primer was incorrect, so PCR was redone today with correct primer just come in.
Results
- No Colonies on plates from this day
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AU CHEM-570 Lab Prep
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