Description
General description
The Rapid DNA Ligation Kit contains all reagents necessary for ligation. No additional reagents or additives are required. The kit can rapidly ligate DNA with either blunt or sticky ends at +15 to +25°C. Depending on the DNA concentration in the reaction, the ligation products will be either circular (if the DNA concentration is low) or concatemeric (if the DNA concentration is high). Ligated DNA is suitable for direct use in transformation experiments.
Note: Electroporation can be performed after ligation in combination with the High Pure® PCR Product Purification Kit.
Application
The Rapid DNA Ligation Kit covalently joins either sticky-ended or blunt-ended DNA fragments, for example, for:
Cloning into plasmid or phage vectors.
Adding linkers to plasmids or other DNAs.
Recircularizing linear DNA.
Biochem/physiol Actions
DNA with either blunt or sticky ends (≤200 ng).
Heat inactivation: 10 min at 65 °C (T4 Ligase)
Inactivation of T4 Ligase is necessary only if the ligation reaction mixture is to be used in experiments other than transformation assays.
Note: Heat inactivation of the ligation reaction mixture before transformation causes the number of transformed colonies to decrease drastically (> factor of 20).
Star activity: Star activity should also be avoided. When double digests are performed with enzymes prone to star activity, consult the Roche Restriction Enzyme Poster (or the Roche Applied Science Catalog) to select the optimal buffer for a given enzyme combination. Whenever possible use Buffer H for double digests.
Features and Benefits
Fast: ligation takes only 5 minutes.
Convenient: ligation is performed at +15°C to +25°C.
Easy: kit contains all buffers and enzymes needed.
Flexible: kit can be used for all common ligation reactions.
Reliable: simple procedure and ready-to-use reagents ensure accuracy and reproducibility.
Packaging
1 kit containing 3 components.
Preparation Note
Incubation time: 5 minutes
Molar ratio of DNA to be ligated: The molar ratio of vector DNA to insert DNA in the standard ligation reaction is normally 1:3. However, when appropriate, you may use other ratios. For example:
If the two DNAs differ greatly in length, try a 1:1 or 1:2 ratio
If the ends of the two DNAs are blunt, try a 1:5 ratio
Other Notes
The new Rapid DNA Dephos & Ligation Kit contains all the reagents of the Rapid DNA Ligation Kit plus our new rAPid Alkaline Phosphatase.
Legal Information
High Pure is a registered trademark of Roche
