Restriction Enzymes and Recombinant DNA

Bacterial cells use restriction enzymes to protect themselves from bacteriophages by cutting viral DNA into smaller, inactive fragments. These enzymes cleave DNA at specific locations, often palindromic sequences.

Many restriction enzymes cut DNA at palindromic sequences, where the sequence reads the same forwards and backwards on opposite strands. These enzymes cleave the DNA, sometimes creating asymmetric cuts.

Asymmetric cuts result in 'sticky ends,' which are single-stranded DNA overhangs that are complementary and can re-anneal. This property is used in creating recombinant DNA.

Restriction enzymes can be used to create recombinant DNA molecules by cutting two different DNA sources with the same enzyme. The complementary sticky ends allow the DNA fragments to combine.

DNA ligase is used to reform the bonds between the DNA fragments, creating a single recombinant DNA molecule made from two different DNA molecules.

Plasmids are used to amplify recombinant DNA. A plasmid is isolated from a bacterial cell, cut with a restriction enzyme, and combined with the target recombinant DNA using DNA ligase.

The recombinant plasmid is inserted into a bacterial cell, which replicates through binary fission, creating many copies of the plasmid containing the recombinant DNA. The recombinant DNA can then be isolated.

The plasmid amplification method has a limitation: there is a size restriction on the recombinant DNA that can be amplified. Very large DNA molecules cannot be effectively amplified using plasmids.