What factors should I consider when selecting restriction enzymes for my experiment?

There are two main factors you need to consider when selecting restriction enzymes for your experiment: 

1. DNA fragment size: Shorter restriction sites (4 base pairs) are more common in genomes, while longer sites (6 to 8 base pairs) are less frequent. To generate larger DNA fragments that are easier to examine by gel electrophoresis, it’s best to use restriction enzymes that recognize and cleave longer restriction sites. 
2. Location and frequency of cuts: Make sure that the restriction enzyme sites occur within the plasmid backbone and flank the gene of interest on either site. This facilitates the creation of compatible sticky ends on both DNA fragments, enabling the construction of recombinant plasmids by ligating the DNA fragments. Carefully check for potential interference with gene or plasmid function by ensuring the chosen enzyme does not cut within the sequence of the gene of interest or within essential plasmid vector components. Cutting within critical regions can disrupt function or alter the encoded products.