The items you’ll need to purchase for your Fiber-Seq experiments depend on whether you’d prefer an out-of-the-box solution or a DIY approach for additional customization. We outline the requirements for both of these options below.
DIY Option
If you are interested in DIY Fiber-seq following our Fiber-Seq protocol, only a few specialized reagents are required in addition to your long-read sequencing workflow. The most critical is the Hia5 enzyme, along with high-quality SAM as its cofactor. Supporting reagents ensure efficient nuclei prep and DNA recovery, enabling reliable Fiber-seq results.
Required Reagents
Hia5 enzyme (EpiCypher 15-1032-8rxn or 15-1032-24rxn)
SAM (cofactor; NEB B9003S recommended) – prepare single-use aliquots to avoid freeze/thaw damage
Nuclei Extraction Buffer (EpiCypher 21-1026)
Monarch® gDNA Extraction Kit (NEB T3010) for DNA recovery
1X PBS (free of calcium and magnesium), any vendor
SDS, any vendor
Molecular biology grade water, any vendor
0.4% Trypan Blue, any vendor
Library Preparation Kit (platform-dependent):
For PacBio HiFi: PacBio SMRTbell Library Prep Kit (e.g. SMRTbell prep kit 3.0)
For Oxford Nanopore Technologies (ONT): ONT Ligation Sequencing Kit (e.g. Ligation Sequencing Kit V14)
Kit Option
If you’d prefer a consolidated solution, our CUTANATM Fiber-seq Kit contains all necessary reagents for nuclei isolation, the 6mA labeling reaction, and gDNA purification. Because the Kit is designed to work with either PacBio or ONT long-read sequencing technology, you will still need to obtain a library preparation kit for your preferred sequencing platform.
Required Reagents
Fiber-Seq Kit (EpiCypher 14-2001-8rxn or 14-2001-24rxn)
1X PBS (free of calcium and magnesium), any vendor
SDS, any vendor
Molecular biology grade water, any vendor
0.4% Trypan Blue, any vendor
Library Preparation Kit (platform-dependent):
For PacBio HiFi: PacBio SMRTbell Library Prep Kit (e.g. SMRTbell prep kit 3.0)
For Oxford Nanopore Technologies (ONT): ONT Ligation Sequencing Kit (e.g. Ligation Sequencing Kit V14)