FAST5

FAST5 is a hierarchical data file format used in nanopore sequencing to store raw electrical signal data and related results. It is built on the HDF5 framework and is commonly associated with Oxford Nanopore Technologies instruments and software. A FAST5 file typically uses the .fast5 extension and serves as a container for sequencing reads, along with metadata about the run and the device.

The internal structure of a FAST5 file is organized into groups and datasets. Each read is stored as a separate group that may contain a raw signal dataset, often labeled Raw, which holds the electrical current measurements collected as a molecule passes through a pore. In addition to the raw signal, FAST5 files can include basecalling results, such as basecall events or basecalled summaries, under nested groups and datasets within the read’s entry. Metadata about the sequencing run, flow cell, chemistry, and instrument is stored as attributes or separate groups at higher levels of the hierarchy.

FAST5 files can be used in two main formats: single-FAST5, which holds one read per file, and multi-FAST5, which aggregates multiple reads into a single file. Multi-FAST5 can improve storage efficiency and I/O performance for large projects, but may require more complex access patterns in analysis pipelines.

In typical workflows, FAST5 serves as the primary source of raw signal data for downstream processing, such as re-basecalling, signal analysis, or alignment. It is read by software libraries and tools used in genomics pipelines, including basecallers and data extraction utilities, making it a foundational component of nanopore data management.