PCR melt curve predictor

Enter a gene, organism, and your known forward and reverse primer sequences (5′→3′). The tool fetches the RefSeq mRNA from NCBI, locates the primers on the template, extracts the amplicon, and predicts a schematic melt curve and Tm (same thermodynamics as the qPCR primer designer).

Use the same buffer assumptions as uMelt: 50 mM [Mono+], 3 mM free Mg2+, SantaLucia & Hicks (2004) nearest neighbours with SL (2004) salt correction. See the melt curve analysis guide for interpreting instrument data.

Leave blank to use RefSeq mRNA from gene lookup. Use NG_ genomic RefSeq if primers were designed on genomic DNA or across splice junctions.

Primary melt peak targets the main derivative peak from SYBR qPCR software (e.g. Applied Biosystems QuantStudio, Bioer QuantGene). Thermodynamic Tm (uMelt-style NN) is also shown and is usually ~3–6 °C higher.