High Quality: Primer3 Input -version 0.4.0-

Unlike older versions, Tm is calculated using the salt-adjusted method by default. Monovalent cation concentration ( PRIMER_SALT_MONOVALENT ) defaults to 50.0 mM.

You must provide the target sequence. Use SEQUENCE for the template. For internal oligos (e.g., hybridization probes), use SEQUENCE_INTERNAL . primer3 input -version 0.4.0-

A more stringent constraint on the 3' end, which is where the polymerase starts extending. Setting this to 3.00 or lower helps prevent dimer formation. Understanding the Output Unlike older versions, Tm is calculated using the

primer3 -version

To avoid repetitive elements or poly-A tracts. Unlike older versions

Mastering Primer3 Input (Version 0.4.0): A Comprehensive Guide to PCR Primer Design