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  • Ligation Protocol for Cloning with Instant Sticky-end Ligase Master Mix (M0370) 

    Protocol

    1. Transfer master mix to ice prior to reaction set up. Mix tube by finger flicking before use.

    2. Combine 20–100 ng of vector* with a 3-fold molar excess of insert and adjust volume to 5 μl with dH2O.

    3. 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.

    4. Use for transformation or store at -20°C.

    5. Do not heat inactivate.

      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.