Product Class: Kit

Quick Ligation™ Kit

Product Introduction

Quick Ligase will ligate these substrates:

dsDNA




Nicked DNA/RNA





 

The Quick Ligation™ Kit enables ligation of cohesive end or blunt end DNA fragments in 5 minutes at room temperature. (25°C )

Catalog # Size Concentration
M2200S 30.0 reactions
M2200L 150.0 reactions

Featured Videos

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    Quick Tips - Do I need a 5' phosphate for ligation?

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    Quick Tips - Electroporation after ligation

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    Quick Tips - Troubleshooting problematic ligation reactions

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    Quick Tip - What is the ideal incubation temperature for ligation?

  • NEBTV_DNA-Modifying-Enzymes-and-Cloning-Techniques_1920

    Quick Tip - When is a long incubation time necessary for ligation?

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    DNA Ligation

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    What are the best conditions for DNA ligation?

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    Why do I need to add PEG to my DNA ligation?

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Product Information

Description

The Quick Ligation Kit enables ligation of cohesive end or blunt end DNA fragments in 5 minutes at room temperature . (25°C )

For details on NEB's quality controls for DNA ligases, visit our Ligase Quality page.
Ligation Time Course:  Ligation Time Course
LITMUS 28i vector was cut with either EcoRV (blunt) or HindIII (cohesive), treated with calf intestinal phosphatase and gel purified. Blunt inserts from a HaeIII digest of ΦX174 DNA and cohesive inserts from a HindIII digest of λ DNA were ligated into the respective vectors at a 3:1 insert:vector ratio using the Quick Ligation Kit. Ligation products were transformed into chemically competent E. coli DH-5α cells and grown overnight on LB-amp plates at 37°C.

1X Quick Ligation Reaction Buffer:
66 mM Tris-HCl
10 mM MgCl2
1 mM Dithiothreitol
1 mM ATP
7.5% Polyethylene glycol (PEG6000)
pH 7.6 @ 25°C

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This product is related to the following categories:
DNA Ligases Products
This product can be used in the following applications:
Cloning Ligation,
Fast Cloning: Accelerate your cloning workflows with reagents from NEB

Kit Components

Kit Components

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration
  • M2200S     -20    
      Quick Ligase M2200SVIAL -20 1 x 0.03 ml Not Applicable
      Quick Ligation Reaction Buffer B2200SVIAL -20 1 x 1 ml 2 X
  • M2200L     -20    
      Quick Ligase M2200LVIAL -20 1 x 0.15 ml Not Applicable
      Quick Ligation Reaction Buffer B2200SVIAL -20 4 x 1 ml 2 X

Properties & Usage

Application Features

Cloning into vectors
Library construction
T/A cloning
Linker ligation
Recirculization of linear DNA

Product Notes

  1. Some of the most critical parameters which should be controlled to ensure successful ligation and transformation are addressed below. 
    Cells: Competent cells can vary by several logs in their competence. Perceived ligation efficiency directly correlates to the competence of the cells used for transformation. Always transform uncut vector as a control for comparison purposes. 
    Electroporation: Electroporation can increase transformation efficiency by several logs. Before using the products of a Quick Ligation reaction for electrotransformation, it is necessary to reduce the PEG concentration. We recommend a spin column purification. 
    DNA: Purified DNA for ligations can be dissolved in dH2O (Milli-Q™ water or equivalent is preferable); TE or other dilute buffers also work well. For optimum ligation, the volume of DNA and insert should be 10 μl before adding 2X Quick Ligation Buffer. For DNA volumes greater than 10 μl, increase the volume of 2X Quick Ligation Buffer such that it remains 50% of the reaction and correspondingly increase the volume of ligase. The overall concentration of vector + insert should be between 1-10 μg/ml for efficient ligation. Insert:vector ratios between 2 and 6 are optimal for single insertions. Ratios below 2:1 result in lower ligation efficiency. Ratios above 6:1 promote multiple inserts. If you are unsure of your DNA concentrations, perform multiple ligations with varying ratios. 
    Time: Most ligations performed using the Quick Ligation™ Kit reach an end point at 5 minutes or less at 25°C. Incubation beyond this time provides no additional benefit. In fact, transformation efficiency starts to decrease after 1 hour and is reduced by up to 75% if the reaction is allowed to go overnight at 25°C. 
    Biology: Some DNA structures, including inverted and tandem repeats, are selected against by E. coli. Some recombinant proteins are not well tolerated by E. coli and can result in poor transformation or small colonies.

Protocols, Manuals & Usage

Protocols

  1. Protocol for the Quick Blunting Kit (E1201)
  2. Transformation Protocol
  3. Quick Ligation Protocol (M2200)

Usage & Guidelines

Application Notes

Tools & Resources

Selection Charts

Web Tools

FAQs & Troubleshooting

FAQs

  1. What factors can cause incomplete ligation and/or transformation using the Quick Ligation Kit?
  2. Why is my transformation not working and what control reactions should be run?
  3. What is the concentration of T4 DNA Ligase provided in the Quick Ligation Kit?
  4. Can conventional T4 DNA Ligase (400,000 units/ml) be used with the Quick Ligation buffer?
  5. How to calculate the molarity of ends?
  6. The protocol for Quick Ligation Kit indicates the reaction should not beheat inactivated. How can I inactivate the ligation activity?

Troubleshooting

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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