Ligation Protocol for Cloning with Instant Sticky-end Ligase Master Mix (M0370)
- Transfer master mix to ice prior to
reaction set up. Mix tube by finger flicking before use.
- Combine 20–100 ng of vector* with a 3-fold
molar excess of insert and adjust volume to 5 μl with
- Add 5 μl of Instant Sticky-end Ligase
Master Mix, mix thoroughly by pipetting up and down 7-10 times, and place on
ice. The sample is now ready to be used for transformation.
- Use for transformation or store at
- Do not heat
Heat inactivation dramatically reduces transformation efficiency.
* In-house testing has demonstrated that maximal transformation efficiency is achieved using between 20–100 ng of vector (sticky) and a corresponding 3-fold molar excess of the insert to be ligated into the vector.