AP BIOLOGY

Detailed explanation-1: -The same restriction enzyme should be used when cutting out a gene of interest and cutting the plasmid vector where the DNA will be inserted so that the sticky ends of both will match up.

Detailed explanation-2: -In molecular cloning a gene of interest can be inserted into a vector, usually a plasmid, by cutting both the vector and the gene (called the insert) with the same enzyme to generate sticky ends and joining the two pieces together to generate a recombinant.

Detailed explanation-3: -Both the plasmid (blue, backbone) and the DNA sequence of interest (green, insert) are cut with restriction enzymes to generate compatible overhangs that allow them to bind. Ligase is used to make bonds between the insert and backbone covalent.

Detailed explanation-4: -How do restriction enzymes work? Like all enzymes, a restriction enzyme works by shape-to-shape matching. When it comes into contact with a DNA sequence with a shape that matches a part of the enzyme, called the recognition site, it wraps around the DNA and causes a break in both strands of the DNA molecule.

Detailed explanation-5: -The sticky ends of the two fragments stick together by complementary base pairing. However, there are still gaps in the sugar-phosphate backbones of the DNA double helix at the junction sites where the gene and plasmid DNA meet. Once they are joined by ligase, the fragments become a single piece of unbroken DNA.