smfprimer

smfprimer designs conversion-aware PCR primer pairs for single-molecule footprinting experiments. Primer3 selects and ranks the pairs; smfprimer adds conversion chemistry, required-interval handling, annotated sequence output, and optional Bowtie 1 specificity assessment.

Choose a workflow

Starting material	Start here
Amplicon or locus FASTA	Amplicon FASTA tutorial
Annotated GenBank or SnapGene file	Annotated-input tutorial
Genome FASTA plus BED/GTF	Genome, BED, and GTF tutorial
Existing forward/reverse primers	Primer evaluation tutorial
Background-genome specificity	Bowtie specificity tutorial

What a design produces

A successful file-based run creates two complementary artifacts:

• A TSV or JSON table containing ranked primer sequences, Primer3 metrics, coordinates, conversion ambiguity, and optional specificity metrics.
• A GenBank file containing the input annotations, required interval, ranked amplicon spans, and strand-aware primer annotations.

Coordinates in CLI arguments, BED input, TSV/JSON output, and the Python API are zero-based and half-open. GTF and GenBank coordinates are converted at the input/output boundary.

Minimal example

smfprimer design fasta docs/examples/amplicon.fa \
  --target-width 10 \
  --product-size 50:100 \
  --workflow conversion \
  --output primers.tsv

This writes primers.tsv and primers.gb.

Note

Primer design is an assay-development aid. Review ambiguity, specificity, product length, and the annotated amplicon before ordering oligos.