SPRING2 Assays and Quality Modes

July 25, 2026 ยท View on GitHub

This guide explains the higher-level data behaviors that affect how SPRING2 stores and restores reads.

Assay Selection

Use --assay / -y to choose an assay explicitly:

spring2 -c --R1 reads.fastq --R2 reads_2.fastq -o reads.sp --assay atac

Supported values are:

  • auto
  • dna
  • rna
  • atac
  • bisulfite
  • sc-rna
  • sc-atac
  • sc-bisulfite

auto is the default. During compression, SPRING2 samples the startup portion of the input and classifies the dataset using assay-specific heuristics and the bundled assay reference.

For technical background on the assay reference, see ../assays/ASSAY_REFERENCE.md.

Cellular Barcode Length

Use --cb-len when single-cell assays store cellular barcodes in the start of R1 instead of a dedicated I1 lane:

spring2 -c --R1 reads.fastq --R2 reads_2.fastq -o reads.sp --assay sc-rna --cb-len 16

Notes:

  • Valid range is 1..64.
  • When --I1 is present, barcode length is inferred from the I1 read length and --cb-len is ignored.

Assay-Aware Compression Behaviors

Depending on assay and confidence, SPRING2 can enable internal transforms that remain reversible for lossless archives.

Examples include:

  • RNA and scRNA: poly-A/T tail stripping when assay detection strongly suggests it is safe
  • ATAC and scATAC: terminal adapter read-through stripping when the overlap is strong enough
  • scRNA and sc-bisulfite: cellular barcode prefix extraction from R1 when no I1 lane is used
  • grouped scRNA with index lanes: trailing index tokens in IDs can be reconstructed from decoded I1/I2 reads

These behaviors are recorded in archive metadata and surfaced by preview mode.

Quality Modes

Use --qmod / -q to choose how quality values are stored.

Lossless

Default mode:

spring2 -c --R1 reads.fastq -o reads.sp -q lossless

This preserves qualities bit-for-bit.

Illumina 8-level Binning

spring2 -c --R1 reads.fastq -o reads.sp -q ill_bin

Binary Thresholding

spring2 -c --R1 reads.fastq -o reads.sp -q binary 20 30 10

This maps qualities to high when >= threshold and to low otherwise.

--strip Modes

Use --strip / -s to intentionally discard archive components:

  • i: identifiers
  • o: original order
  • q: quality scores

Example:

spring2 -c --R1 reads.fastq --R2 reads_2.fastq -o reads.sp --strip io

Be careful: stripping changes what can be faithfully restored later.

When to Prefer Explicit Assay Selection

Prefer --assay over auto when:

  • you already know the assay and want deterministic behavior
  • startup samples are atypical or low quality
  • you are benchmarking and want consistent cross-run settings

See Also