Team:Brown-Stanford/Lab/Protocol/T4Ligation

T4 ligation

(Taken from openwetware.org; this protocol was edited for amplification of Part BBa_K656013)

Reagents for 35µL reaction

• 2.9 µL nuclease free water
• 3.5 µL 10x T4 ligation buffer
• 24 µL insert (BBa_K656013, 10 ng/µL)
• 2.85 µL vector (pSCB3, 14.6ng/µL)
• 1.75 µL T4 ligase

Treatment

1. Add all reagents in order
2. Allow to incubate for 2 hours at room temperature
3. Denature ligase at 65˚C, store at 4˚ or -20˚