Question

What criteria do restriction enzymes have to meet when deciding which to choose in order to clone a specific gene into the multiple cloning site of a plasmid (e.g. pUC18)?

Answer 1

The main criteria restriction enzymes must meet for the application in cloning a specific gene into the multiple cloning site of a plasmid are:

- Flank the insert, but does not cut within the insert;
- Is in the desired location in the recipient plasmid;
- Results in the insert being in the correct orientation in the recipient plasmid;
- Is in the frame with tags or fusion proteins in the recipient plasmid.