Adaptor Ligation (E7370)

 This caution sign signifies a step in the protocol that has multiple paths leading to the same end point but is dependent on a user variable, like the amount of input DNA.


  1. Add the following components directly to the End Prep reaction mixture and mix well: 
    Blunt/TA Ligase Master Mix   15 μl 
    NEBNext Adaptor for Illumina*   2.5 μl 
    Ligation Enhancer   1 μl 
    Total volume   83.5 μl

    *The NEBNext adaptor is provided in NEBNext Singleplex (NEB #E7350) or Multiplex (NEB #E7335 #E7500) Oligos for Illumina.

  2. Mix by pipetting followed by a quick spin to collect all liquid from the sides of the tube.

  3. Incubate at 20°C for 15 minutes in a thermal cycler.

  4. Add 3 μl of USER™ enzyme to the ligation mixture from step 3.

  5. Mix well and incubate at 37°C for 15 minutes.