15–17	μL Adaptor-ligated CUT&RUN DNA fragments (from Step 19)
25 μL	NEBNext® Ultra™ II Q5® Master Mix
2.5 μL	Index Primer for Illumina*
2.5 μL	Universal PCR Primer for Illumina
Up to 50 μL	Nuclease-free H2O

^*

Index primer should be different for each sample that will be pooled together in one sequencing lane. Keep careful notes of which index primer is added to each reaction.