Relative activity of restriction enzymes in universal and basal buffers
Our restriction enzymes are supplied with an optimal universal buffer (one of five universal buffers; indicated in blue in the table below). The relative activity in each of the other universal buffers is normalized to the optimal buffer, where the activity of each enzyme in the optimal buffer is expressed as 100%. Values in ( ) indicate buffers that are likely to be affected by star activity. In order to avoid these effects, use of buffers highlighted in blue or pink is recommended.
A few specific enzymes (AccIII, BalI, BcnI, BglI, Bpu1102I, Cfr10I, Eco52I, NruI, PshBI, SnaBI, SspI, TaqI, and VpaK11BI), are each supplied with a basal buffer specialized for the particular enzyme. The compositions of these basal buffers vary depending on the enzyme.
For information on double digestion, see the universal buffers for double digestion with restriction enzymes page.
The tables below list relative activities of restriction enzymes in the universal buffers and instructions for use and compositions of buffers.
Blue: buffer supplied with the restriction enzyme
Pink: alternative buffer recommended for use
*+0.01% BSA → 100% AflII, EcoO65I, FokI, Hin1I, MunI, NcoI, PvuI, SplI, Sse8387I, XbaI
**+0.01% BSA + 01% Triton X-100 → 100% NotI
*** The compositions of the basal buffers are enzyme-specific.
Instructions for the use of loading buffers
All restriction enzymes are supplied with a 10X loading buffer (1 ml) containing 1% SDS, 50% glycerol, and 0.05% bromophenol blue. Add >1/10 volume of 10X loading buffer to stop the digestion reaction, and run on an agarose gel.
Note: SDS may precipitate during storage at room temperature. If there is a precipitate, dissolve in a warm water bath before use.
Instructions for the use of universal buffers
Universal buffers are provided at a 10X concentration. Please add 1/10 the volume of the reaction mixture. Since 10X T4 buffer does not contain BSA, be sure to add BSA to a final concentration of 0.01%. Some restriction enzymes are supposed to exhibit 100% activity when BSA or Triton X-100 is added to the reaction system. Since BSA and Triton X-100 are supplied with enzymes at a 10X concentration (0.1%), be sure to add 1/10 the volume of the reaction mixture to the buffer before starting the reaction.
Compositions of provided universal buffers and other reagents
| Universal buffer | Composition |
| 10X L | 100 mM Tris-HCl (pH 7.5) |
| 100 mM MgCl2 | |
| 10 mM Dithiothreitol | |
| 10X M | 100 mM Tris-HCl (pH 7.5) |
| 100 mM MgCl2 | |
| 10 mM Dithiothreitol | |
| 500 mM NaCl | |
| 10X H | 500 mM Tris-HCl (pH 7.5) |
| 100 mM MgCl2 | |
| 10 mM Dithiothreitol | |
| 1,000 mM NaCl | |
| 10X K | 200 mM Tris-HCl (pH 8.5) |
| 100 mM MgCl2 | |
| 10 mM Dithiothreitol | |
| 1,000 mM KCl | |
| 10X T (BSA-free) |
330 mM Tris-acetate (pH 7.9) |
| 100 mM Mg-acetate | |
| 5 mM Dithiothreitol | |
| 660 mM K-acetate | |
| 0.1% BSA dissolved in sterile water | |
| 0.1% Triton X-100 | |
In-Fusion applications and tech notes
View application data on how In-Fusion technology performs for all of your cloning needs.
In‑Fusion Cloning tips and FAQs
Learn more about In‑Fusion Cloning, including applications, tips, primer design, and vector and insert requirements.
Restriction enzyme finder
Choose your restriction enzyme by overhang type, recognition site size, or alphabetical listing.
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