For suggestions for optimal cleavage of a restriction site occurring at the end of a DNA molecule please refer to the video above.
Proximity to the end of a DNA molecule can influence restriction enzyme digestion. As a general rule, adding 6 nucleotides between the end of the primer and the 5' end of the recognition site typically ensures efficient cleavage. It is important to make sure that the added bases are not palindromic and won't form primer dimers. For more specific experimental data detailing requirements for individual restriction enzymes, select the link.
