Cracking the AP Biology Exam

6

Molecular Genetics

RECOMBINANT DNA

Recombinant DNA is a hybrid of DNA from two or more sources. For example, DNA with a gene of interest from eukaryotic organisms can be transferred into a bacterial genome. The branch of technology that produces new organisms or products by transferring genes between cells is called genetic engineering.

How does this procedure work? A restriction enzyme recognizes a short, DNA sequence, a restriction site, and cuts both DNA strands at recognition sequences. The resulting double-stranded restriction fragments have at least one single-stranded end called a sticky end. These short extensions allow hydrogen-bonded base pairs between the sticky ends of a plasmid, a cloning vector, and the DNA molecule of interest cut with the same restrictive enzyme. When DNA ligase permanently seals the two fragments a recombinant DNA molecule is produced.

The recombinant DNA is then inserted into a bacterium by transformation. The host cell is grown in a culture to form a clone of cells containing the “cloned” gene of interest. For example, this technology is used to produce insulin when inserted into E. coli.

The DNA fragments can be separated according to their molecular weight using gel electrophoresis. Because DNA and RNA are negatively charged, they migrate through the gel toward the positive pole of the electrical field. The smaller the fragments, the faster they move through the gel. Restriction enzymes are also used to create a molecular fingerprint. When restriction fragments between individuals of the same species are compared, the fragments differ in length because of polymorphisms, which are slight differences in DNA sequences. These fragments are calledrestriction fragment length polymorphisms, or RFLPs. In DNA fingerprinting, RFLPs produced from DNA left at a crime scene are compared to RFLPs from the DNA of suspects.