1. What is a Ligation Molar Ratio Calculator?

Definition: This calculator determines the molar ratio between DNA vector and insert for optimal ligation reactions.

Purpose: It helps molecular biologists achieve efficient ligation when cloning DNA fragments into vectors.

2. How Does the Calculator Work?

The calculator uses the formula:

Where:

• \( R \) — Molar ratio (dimensionless)
• \( n_D \) — Moles of DNA vector (mol)
• \( n_I \) — Moles of insert DNA (mol)

Explanation: The ratio of DNA vector moles to insert moles determines the optimal stoichiometry for ligation reactions.

3. Importance of Molar Ratio in Ligation

Details: Proper molar ratios (typically 1:1 to 1:3 vector:insert) minimize empty vector backgrounds and maximize correct recombinants.

4. Using the Calculator

Tips: Enter the moles of DNA vector and moles of insert. Both values must be > 0. For best results, use values in the picomole to nanomole range.

5. Frequently Asked Questions (FAQ)

Q1: What's the ideal molar ratio for ligation?
A: Most protocols recommend 1:1 to 1:3 (vector:insert), with 1:2 being common for standard cloning.

Q2: How do I convert ng of DNA to moles?
A: Use the formula: moles = (mass in g)/(660 g/mol × length in bp). We recommend using NEB's "DNA Molarity Calculator".

Q3: Why is molar ratio important?
A: Proper ratios ensure efficient ligation while minimizing self-ligation of vector or multimerization of insert.

Q4: When would I use different ratios?
A: Higher insert ratios (1:3) may help with difficult ligations or short inserts, while 1:1 may work better for large inserts.

Q5: Does this work for blunt-end ligations?
A: Yes, though blunt-end ligations often require higher insert ratios (up to 1:10) due to lower efficiency.