Useful tips

What is digest buffer?

What is digest buffer?

$ 481.00. Differex™ Digestion Buffer is used in differential extractions of samples that contain a mixture of epithelial and sperm cells. A digestion combination of this buffer and Proteinase K can be used to selectively lyse epithelial cells while leaving sperm cells intact.

What does restriction buffer do?

A restriction digest is a procedure used in molecular biology to prepare DNA for analysis or other processing. These enzymes are called restriction endonucleases or restriction enzymes, and they are able to cleave DNA molecules at the positions at which particular short sequences of bases are present.

Why is my restriction digest not working?

If there are no issues with digesting the control DNA, there may be something else wrong with your reaction set up. Double check the optimal reaction temperature for the enzymes being used, and control for evaporation during incubation as an increased salt concentration in the buffer can inhibit enzyme performance.

What are the ways to stop a restriction digestion reaction?

Restriction Enzyme Digest Protocol

  1. Add components to a clean tube in the order shown:
  2. Incubate the reaction at digestion temperature (usually 37 °C) for 1 hour.
  3. Stop the digestion by heat inactivation (65 °C for 15 minutes) or addition of 10 mM final concentration EDTA.

Is heat inactivation of restriction enzymes necessary?

FAQ: Is it necessary to inactivate restriction enzymes after vector digestion? Inactivation of restriction endonucleases is generally not necessary, but in some cases it might increase the transformation efficiency.

How many restriction enzymes are in Neb buffer?

NEB’s restriction enzyme buffer system makes your restriction digests easy and convenient. We are able to offer >215 restriction enzymes that cut in a single buffer, CutSmart ® .

What kind of nebuffer is used for Double Digest?

Double digests with NEB’s restriction enzymes can be set up in CutSmart Buffer. Otherwise, choose an NEBuffer that results in the most activity for both enzymes. If star activity is a concern, consider using one of our High Fidelity (HF®) enzymes. Set up reaction according to recommended protocol.

Is there a Double Digest protocol for neb?

Please note that NEBcloner will also provide detailed double digest protocols using this enzyme. Additional information on performing digests using restriction enzymes can be found in our reference article: Optimizing Restriction Endonuclease Reactions.

How are restriction enzymes digested in PCR buffers?

For convenience, restriction enzyme digestion can be performed directly in the PCR mix without any purification of the DNA. This table summarizes the percent activity of restriction enzymes on the DNA in the Taq, Phusion® or Q5® PCR mixes described below.