Protocol 5: UltraShear fragmentation of tissue genomic DNA

Why UltraShear is an enzyme reagent—not a machine—and what its fragment distribution contributes

First, a correction

NEBNext UltraShear (M7634) is an enzymatic fragmentation reagent, not an instrument. A thermal cycler controls incubation. The formulation is designed to fragment DNA upstream of short-read library preparation while retaining methylation marks.

Input	Reaction	Output	Immediate data
purified tissue genomic DNA	timed UltraShear incubation and heat inactivation	fragmented tissue DNA	fragment-size trace, concentration, yield

The NEB manual describes 5–250 ng purified gDNA and time-dependent fragmentation at 37°C followed by heat inactivation; the exact study conditions must be fixed during validation. The manual warns that the coupled EM-seq protocol differs from standard standalone paths.

Why only tissue?

Tissue genomic DNA needs controlled fragmentation for Illumina library construction. Plasma cfDNA is already fragmented in vivo; deliberately re-shearing it would erase or distort the native fragmentomic signal.

QC gate

Review the full fragment distribution—not only its mean—plus input mass, reaction time, buffer, lot, replicate, and recovery. Fragmentation must be balanced across groups. Tissue insert length is a preparation characteristic, not a nucleosome footprint.

Official UltraShear product page → · Next: EM-seq library preparation →