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Use the NEB Tm Calculator to estimate an appropriate annealing temperature when using NEB PCR products.
Use this tool when designing PCR reaction protocols to help determine the optimal annealing temperature for your amplicon. Simply input your DNA polymerase, primer concentration and your primer sequence and the Tm Calculator will guide you to successful reaction conditions.
Understand the expected properties of your oligos before you order them. Calculator for GC content, melting temperature (T m), molecular weight, extinction coefficient, µg/OD, nmol/OD, and more. Identify secondary structure potential. Minimize dimerization. Use NCBI BLAST™.
The calculator calculates recommended T m (melting temperature) of primers and PCR annealing temperature based on the primer pair sequence, primer concentration, and DNA polymerase used in PCR. The calculator also calculates the primer length, percentage of GC content, molecular weight, and extinction coefficient.
8 lis 2021 · PCR primers are interdependent with your sample type, target, reagents, and thermodynamic conditions. Establishing your PCR objective can guide your primer design and enzyme choices. It’s a good idea to review parameters and understand why primer design is such a critical, dependent variable in PCR.
Whether you are creating an assembly that incorporates multiple fragments, conducting your entire PCR at a single temperature, or incorporating intentionally mismatched nucleotides, careful primer design can set you up for success before you even pick up your pipette.
21 paź 2013 · The free, online IDT OligoAnalyzer Tool allows researchers to calculate T m for mismatched sequences. You can also use the program to determine probe locations that help enable better mismatch discrimination.
