Vector Ligation Ratio Calculator

Calculate optimal vector to insert molar ratios for successful DNA ligation and cloning experiments

kb
kb
ng

Insert DNA Required at Different Molar Ratios

Molar Ratio
(Insert:Vector)		Insert Mass
(ng)		Total DNA
(ng)		Recommended Use
1:1 16.7 66.7 Blunt or single digest
2:1 33.3 83.3 Standard cloning
3:1 50.0 100.0 Standard cloning (recommended)
4:1 66.7 116.7 Difficult ligations
5:1 83.3 133.3 Small inserts or difficult ligations
6:1 100.0 150.0 Very small inserts
7:1 116.7 166.7 Very small inserts
8:1 133.3 183.3 Extremely small inserts
10:1 166.7 216.7 Extremely small inserts
Insert mass (ng) = [Vector mass (ng) × Insert length (kb) × Molar ratio] / Vector length (kb)

What is Vector Ligation?

Vector ligation is a molecular cloning technique where a DNA insert is joined to a vector (plasmid) using DNA ligase enzyme. The molar ratio between vector and insert is crucial for successful ligation.

How to Use This Calculator

  1. Enter your vector length in base pairs
  2. Enter your insert length in base pairs
  3. Enter the amount of vector DNA
  4. Select desired molar ratio
  5. View calculated insert amount needed

Optimal Ratios

  • 1:3 - Most common ratio for routine cloning
  • 1:1 - For blunt-end ligations
  • 1:5 - For difficult ligations or small inserts
  • Higher ratios may reduce self-ligation

Tips for Success

  • Use fresh ligase and buffer
  • Dephosphorylate vector to prevent self-ligation
  • Keep total DNA concentration between 1-10 ng/µL
  • Incubate at 16°C overnight or room temperature for 1 hour

Common Applications

  • Gene cloning and expression
  • Subcloning DNA fragments
  • Creating fusion proteins
  • Library construction
  • Site-directed mutagenesis

FAQ

Q: Why is molar ratio important?
A: Proper molar ratio ensures optimal insert incorporation and minimizes vector self-ligation.

Q: What if my ligation fails?
A: Try different ratios, check DNA quality, ensure compatible ends, and verify ligase activity.