PCR Primer & Pairwise Dimer Thermodynamic Analyzer

• Stage 1 (Single Oligo): Evaluates length, GC%, Tm, 5'-G check, nucleotide runs, 3'-CG clamp, hairpin loops, and homodimers.
• Stage 2 (Pairwise Matrix): Evaluates all 2-combinations with replacement (homodimers & heterodimers) for cross-hybridization risk.
• Degenerate Bases: Automatically expands ambiguous IUPAC nucleotides (W, R, Y, S, K, M, B, D, H, V, N) into consecutive canonical variants.
• Temperature Controls: Distinct baseline settings for structural ΔG simulation (temp_c) and PCR annealing adjustments (annealing_temp_c).

📖 Read Complete Instructions & Guidelines

Your input file must be a comma-separated values (.csv) file structured with exactly two columns:

• Column 1: Primer identifier with the exact heading Name
• Column 2: 5' to 3' nucleotide sequence with the exact heading Sequence

Example Format: