NEB ChIP-Seq Library Prep Master Mix Set for Illumina User Manual

About this manual

This manual provides step-by-step protocols for preparing DNA libraries for next-generation sequencing of chromatin-immunoprecipitated DNA using NEBNext reagents. It includes detailed procedures for end repair, dA-tailing, adaptor ligation, size selection, and PCR enrichment, with specific instructions for using NEBNext singleplex or multiplex oligos. The document also covers cleanup steps using AMPure XP beads and storage conditions for reagents.

The master mix set contains pre-formulated enzymes and buffers, including Klenow Fragment, Quick T4 DNA Ligase, and Phusion High-Fidelity PCR Master Mix, all stored at –20°C. The manual emphasizes lot control and functional validation of the reagents for consistent library preparation results.

What's inside

• Applications
• Protocol for Use with NEBNext Singleplex or Multiplex Oligos for Illumina
• Protocol for Use with End User Supplied Primers and Adaptors
• NEBNext End Repair Enzyme Mix
• NEBNext End Repair Reaction Buffer
• Klenow Fragment (3´→5´ exo–)
• NEBNext dA-Tailing Reaction Buffer
• Quick T4 DNA Ligase
• NEBNext Quick Ligation Reaction Buffer
• Phusion® High-Fidelity PCR Master Mix with HF Buffer

Frequently asked questions

What is the storage condition for the reagents in the master mix set?

All reagents should be stored at –20°C.

How many reactions can be prepared with the NEB #E6240S and NEB #E6240L kits?

The NEB #E6240S kit is sufficient for 12 reactions, and the NEB #E6240L kit for 60 reactions.

What is the recommended DNA starting material for the protocol?

10 ng of chromatin-immunoprecipitated (ChIP) qPCR verified or control DNA, in ≤ 40 µl of water or elution buffer.