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Timeline

Week 26

- Preparing for take-off

• Inventory of the lab supplies
• Pouring LB Agar plates
• Amplification of the pET-Duet-1 vector
• Assessment of the safety requirements
• Preparing stocks for antibiotics, glycerol, LB & MilliQ

Week 27

- The Clone Wars

Gibson Assembly:

• Linearizing pET-Duet-1 for Gibson Assembly
• Our first Gibson Assembly!

Traditional cloning:

• Amplification of the pET-Duet-1 vector
• Nde1 & Kpn1 digestion of the pET-Duet-1 vector (MCS-2) for traditional cloning

Week 28

- Et tu, pET-Duet?

Gibson Assembly:

• Linearizing pET-Duet-1 for Gibson Assembly and debugging
• Amplification of the inserts for traditional cloning
• Xbal & Pstl digestion of the pET-Duet-1 vector (MCS-1) for traditional cloning

Traditional cloning:

• Xba1 & Pst1 digestion of the inserts (MCS-1) for traditional cloning
• Nde1 & Kpn1 digestion of the inserts (MCS-2) for traditional cloning
• Ligation