NCBI Primer-BLAST

NCBI Primer-BLAST is a free tool from the National Center for Biotechnology Information that integrates the Primer3 primer design engine with the BLAST sequence alignment algorithm. It designs PCR primers and simultaneously checks their specificity against a comprehensive nucleotide database, ensuring that primers will only amplify the intended target and not produce off-target products. Users can input accession numbers, GI numbers, or FASTA sequences, specify exon/intron junction spanning for mRNA targets, and set organism-specific specificity filters. It is the gold-standard tool for designing highly specific PCR primers for gene expression, cloning, genotyping, and diagnostic applications.