Off-target Detection

Feature documentation: Off-target Detection

v0.3.0 - BLAST-based specificity analysis.

Overview

PrimerLab checks primer specificity using BLAST alignment against reference databases.

Features

• Dual Backend: BLAST+ or Biopython fallback
• Parallel Processing: Multi-threaded for speed (v0.3.2)
• Caching: Skip redundant alignments
• Scoring: A-F grades based on off-target count

CLI Usage

Standalone

primerlab blast -p primers.json -d genome.fasta

Integrated with Design

primerlab run pcr --config my_config.yaml --blast --blast-db genome.fasta

Config-based

# my_config.yaml
offtarget:
  enabled: true
  database: /path/to/genome.fasta
  mode: auto  # auto, blast, biopython

Scoring System

Grade	Score	Off-targets	Risk
A	90-100	0-2	Minimal
B	80-89	3-5	Low
C	70-79	6-10	Moderate
D	60-69	11-20	High
F	<60	>20	Severe

Output

{
  "forward": {
    "offtargets": 2,
    "score": 95.0
  },
  "reverse": {
    "offtargets": 1,
    "score": 97.5
  },
  "combined_score": 96.25,
  "grade": "A"
}

Backend Selection

Mode	Description
auto	BLAST+ if available, else Biopython
blast	Force BLAST+ (faster, requires install)
biopython	Pure Python (slower, always available)

See Also

• BLAST Command - Detailed CLI options
• In-silico PCR - Template validation