Use this free online primer binding checker and primer map to find exact primer binding sites on linear or circular DNA, review orientation and additional matches within the entered template, detect 5' overhangs, and calculate expected amplicon sizes.
Useful Checks
You can add multiple primer pairs and check them against the same template in one run.
Use the primer map to compare the expected site with additional exact matches elsewhere in the entered template. This is not a genome-wide off-target search.
After checking binding, the current template can be opened in Primer Designer or Restriction Site Analyzer for the next primer-design or restriction-check step.
Primer Binding Checker can receive primer/template handoffs from Primer Designer and Sanger Primer Designer, so design and validation stay in one browser workflow.
Minimum Inputs Needed
Template DNA sequence.
One forward primer.
One reverse primer.
Optional Settings
Template topology
Use linear for fragments. Use circular for plasmids. In circular mode, the coordinate origin means base 1 / the start of the entered sequence, not the biological ori.
Add primer pair
Use this to check more than one primer pair on the same template.
Minimum binding length, bp
This is the shortest exact 3' match accepted as binding. Keep the default unless you are troubleshooting.
Max matches shown per primer
Limits long lists when a primer binds many places.
Open in other tools
Send the same template to Primer Designer or Restriction Site Analyzer.
How To Use
Paste or upload the template.
Choose linear or circular topology.
Enter primers in 5' to 3' direction.
Add more primer pairs if needed.
Click Check primer binding to build the primer map and specificity summary.
Understanding The Results
The quick result tells you whether a product is expected.
The primer map shows where the primers bind.
5' overhangs are shown separately from the template-binding part.
Multiple binding sites are a warning for poor primer specificity or nonspecific PCR.
The template amplicon size spans the selected binding sites. Final PCR product size also includes detected 5' overhang bases.
The summary and detailed map can be copied for records or troubleshooting.
Accepted Input Formats
Template: raw DNA sequence, FASTA, or GenBank text with an ORIGIN sequence section.
Primers: A/T/G/C DNA sequence, written 5' to 3'.
FASTA headers and GenBank annotations are ignored. Multiple FASTA records are concatenated, so analyze separate templates one at a time.
.dna / SnapGene binary files are not supported. Export as FASTA or GenBank.
Assumptions And Limitations
The tool requires exact binding in the accepted 3' binding region. It does not allow mismatches, degenerate primer bases, or inosine.
Minimum binding length can identify an exact 3' binding suffix while treating unmatched 5' sequence as an overhang; it is not mismatch tolerance.
The tool does not calculate primer Tm or thermodynamic binding strength.
A result with no binding warnings does not guarantee PCR success.
Use the result as a screening check, then confirm with your experiment conditions.
The browser tool accepts sequence files smaller than 2 MB and templates up to 500,000 bp.
Example
Paste a plasmid sequence, enter your forward and reverse primers, choose circular topology, and check that each primer binds once with the expected product size.
Use note: These tools are for research and educational planning. Check important calculations and sequence designs before ordering reagents or running experiments.