Tm Calculator

The DNA/RNA Melting Temperature (Tm) Calculator allows users to estimate the melting temperature of nucleic acid duplexes by inputting sequence length, GC content, salt concentration, and oligonucleotide type to obtain a basic, salt-adjusted, and final adjusted Tm.

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Step-by-Step Guide to Using the DNA/RNA Melting Temperature (Tm) Calculator

This guide will help you navigate and use the DNA/RNA Melting Temperature (Tm) Calculator efficiently. Follow these steps to calculate the melting temperature of your oligonucleotide sequences.

Step 1: Input Sequence Length

  • Locate the Sequence Length (bp) field.
  • Enter the total number of base pairs for the sequence you are analyzing. This value should be a positive number between 1 and 1000.
  • This input is mandatory, so ensure it is entered correctly to proceed with the calculation.

Step 2: Enter GC Content

  • Find the GC Content (%) field.
  • Input the percentage of guanine and cytosine bases in your sequence. The valid range is from 0% to 100%, and you can use increments of 0.1% for precision.
  • Make sure this field is completed as it is essential for calculating the Tm.

Step 3: Specify Salt Concentration

  • Identify the Salt Concentration [Na+] (mM) field.
  • Enter the sodium ion concentration in millimolar within the range of 0 to 1000 mM. It’s important to provide an accurate concentration as it impacts the Tm calculation.
  • This is a required input, so ensure it’s filled in properly.

Step 4: Choose Oligonucleotide Type

  • Locate the Oligonucleotide Type dropdown menu.
  • Select one of the following options based on your sequence type: DNA/DNA, RNA/RNA, or DNA/RNA Hybrid.
  • This selection influences the final Tm adjustment, so choose the appropriate type carefully.

Step 5: View Results

  • Once all the fields are correctly filled, the calculator will display the calculated melting temperatures.
  • First, the Basic Tm (°C) will be shown, calculated using the formula: 69.3 + (0.41 * GC content) – (650 / sequence length).
  • Next, the Salt-Adjusted Tm (°C) appears, adjusting the basic Tm by the formula: basic Tm + (12.5 * log10(salt concentration/1000)).
  • Finally, the Final Adjusted Tm (°C) is shown, further adjusting the Tm based on the oligonucleotide type: Tm for RNA is increased by 2°C, for hybrids by 1°C, and for DNA it remains the same as the salt-adjusted Tm.

Ensure all inputs are accurate for the most reliable melting temperature results. You can adjust input values as needed and observe how they influence the Tm outcomes.