NEBcutter® V3.0 – Visualize a restriction digest on a virtual gel
This tutorial explains how to use NEBcutter V3.0 to visualize a restriction enzyme digest on various gel systems by inputting your electrophoresis parameters.
Script
If you have a sequence you would like to analyze, you can paste it in the DNA sequence box. You can also select from frequently used plasmids and phage genomes. Or select from a previous project.
Let’s visualize a virtual digest of the Lambda Phage genome. Click on the Viral & Phage option and select Lambda NEB from the menu. Lambda DNA is linear, so leave circular unchecked. Click “Submit”. The resulting image only indicates enzymes that cleave once. Since PaqCI cleaves more than once, we need to use the NEBcutter Custom Digest feature. Click “custom digest”.
You can search for a specific enzyme by name or scroll through the list to find it. Select PaqCI from the list. Click “digest”. NEBcutter displays a map of the sequence with PaqCI sites displayed. To visualize a gel of this custom digest, click “Gel”.
You can input the length of your gel to better match your electrophoresis conditions. If you are using other gel systems, you can input your electrophoresis parameters corresponding to the conditions used with your electrophoresis setup. You can select the type of electrophoresis gel that you are using with your samples, as well as the percentage.
You can include a DNA ladder marker on your virtual gel using the drop-down selection menu. Let’s use the NEB 1kb ladder as our marker. You can now see the expected band pattern for a PaqCI digest of Lambda DNA.
You can find NEBcutter at nebcutter.neb.com.
For more information on restriction enzymes and how-to videos visit www.neb.com
Related Videos
-
Standard Protocol for Restriction Enzyme Digests -
Restriction Enzyme Digestion Problem: DNA Smear on Agarose Gel -
Restriction Enzyme Digest Problem: Too Many DNA Bands
