ATAC-seq¶

ATAC runs remove duplicates by default, filter at MAPQ 30, report mitochondrial read statistics, and use replicate-aware IDR consensus for narrow peaks.

omnomnomics atac \
  -i EXPERIMENT \
  -g GRCh38 \
  -m metadata.tsv \
  --sample-name condition,donor,replicate \
  --sample-type condition

The workflow continues after common step 9 with peak calling, peak QC, pre-DE interpretation, counting, differential chromatin analysis, and post-DE interpretation (steps 10-15).

Peak construction¶

--narrow-peak-strategy idr is the default. MACS3 calls each biological replicate separately, and IDR derives reproducible group-level peaks from the replicate calls. --narrow-peak-strategy macs3 instead pools each group and can choose MACS3 parameters using the optimizer below.

Control	Behavior
--idr-mode basic	run pairwise IDR on the true biological replicates
--idr-mode encode	add pooled and per-replicate pseudoreplicate diagnostics to the true-replicate analysis
--idr-pairing-policy all_pairs	compare every replicate pair; most complete for groups with more than two replicates
--idr-pairing-policy anchor_vs_all	compare the first replicate with each remaining replicate; fewer jobs for large groups

With the MACS3 strategy, --atac-peak-opt-mode none makes one fixed call at q=0.01, shift=-100, and extension size=200. fast compares that call with a shift=-75/extension=150 alternative. full evaluates six combinations spanning q=0.01 or 0.05 and shift/extension pairs of -100/200, -75/150, and -50/100.

For each group, the optimizer favors high FRiP and agreement between replicate peak sets, penalizes blacklist overlap, and weakly favors a median peak width near 250 bp. It writes all candidate scores and the selected parameters under atac_peak_call_optimization/.

SPP quality gate¶

--spp-gate none disables decisions based on NSC/RSC. warn records threshold failures without changing downstream samples. drop excludes flagged samples from counting and differential analysis. strict stops the workflow if any sample fails the configured thresholds.

Install the IDR helper for the default strategy and the SPP helper when NSC/RSC metrics or SPP gate decisions are required.