1. What is Ligation Molar Ratio?

The Ligation Molar Ratio (MR) is the ratio of insert DNA moles to vector DNA moles in a ligation reaction. This ratio is crucial for efficient cloning as it affects the likelihood of getting the desired recombinant product.

2. How Does the Calculator Work?

The calculator uses the simple formula:

Where:

• \( MR \) — Molar ratio (water irrelevant)
• \( MI \) — Insert moles
• \( MV \) — Vector moles

Explanation: The ratio determines the stoichiometry of insert to vector in the ligation reaction, with optimal ratios typically between 1:1 and 3:1 (insert:vector).

3. Importance of Molar Ratio in Ligation

Details: Proper molar ratios are essential for efficient ligation. Too much insert can lead to multiple inserts, while too little can result in empty vectors. The water volume is irrelevant as it's the molar ratio that matters.

4. Using the Calculator

Tips: Enter the amount of insert and vector in moles. The calculator will compute the molar ratio. Typical ligations use 50-100 ng of vector DNA.

5. Frequently Asked Questions (FAQ)

Q1: What is the ideal molar ratio for ligation?
A: For most applications, a 3:1 insert:vector ratio works well, though some protocols recommend 1:1 to 5:1 depending on insert size.

Q2: Why is water irrelevant in the calculation?
A: The reaction efficiency depends on the molar ratio of reactants, not their absolute concentrations in the reaction mix.

Q3: How do I convert ng of DNA to moles?
A: Use the formula: moles = (mass in g) / (length in bp × 660 g/mol/bp). Our calculator focuses on the ratio once you have mole values.

Q4: Does insert size affect the optimal ratio?
A: Yes, larger inserts may require higher ratios (up to 10:1) to compensate for lower ligation efficiency.

Q5: Can I use this for blunt-end ligations?
A: Yes, but blunt-end ligations typically require higher insert:vector ratios (5:1 to 10:1) due to lower efficiency.