Cloning Strategy

CD79A Linker Optimization

Antibody Choices 

Sunday's To Do's

• Take out plates
• Miniprep (if you can find 2mL tubes...)
  • pENTR HyHEL10 Heavy
  • hEF1a:CD79B-9T15-G4 
• Pick colonies
  • Plated 8/8
    • A-15T9
    • TRE:Syk-3-TEVp
    • TRE:Syk-6-TEVp
    • TRE:Syk-9-TEVp
    • TRE:Syk-12-TEVp
    • B-3T15
    • B-9T9
    • TRE:Syk-15-tTEVp
  • Streaked or restreaked 8/8
    • A-3T15
    • B-15T15
    • B-15T9
    • B-6T12
  • Plated previously
    • A-9T15
• Start midiprep cultures
  • B3T3
  • B3T9
  • B6T6
  • B9T3
  • B9T6
  • B12T6
  • B15T3
• Send for sequencing
  • Minipreps (if they get done)
    • pENTR HyHEL10 Heavy
    • hEF1a:CD79B-9T15-G4 
  • CD79B linkers (#1 from before)
    • B9T12
    • B6T9
      

Saturday's To Do's

• Kill LR (TRE:Syk-15-tTEVp)
• Take out plates + colonies 
• Transform
  • hEF1a:CD79A-15T9-G4 (because even though we midi'ed it, sequencing was bad)
  • TRE:Syk-linker-TEVp (3, 6, 9, 12)
  • hEF1a:CD79B-3T15-G4
  • hEF1a:CD79B-9T9-G4
  • TRE:Syk-15-tTEVp
• Streaked out
  • A-3T15
  • B-15T15
  • B-15T9
  • B-6T12

Sequencing account information:
^^^Email: bcrpeeps@mit.edu

^^^Pass: grapefruit

Non-Lab To Do's 

• Cytometry time 
• Order Cumate 
• Chicken lysozyme, gp120
• Antibodies for antigen binding detection

Waiting on primers to do list:

Higher Level To Do's:

• Decide on other antigen-variable regions we want to test  
• Is gantenerumab proprietary and can we actually publish with it?

THINGS TO NOT FORGET TO DO:

• SYK-15tTEVP golden gate when we figure out what's wrong with the AarI
• 3T3 MIDI