IV.

Gene Cloning and Hybridization

Although the major advances described above were made in the 1950s and 1960s, the explosion in molecular biology began in the 1970s with the development of techniques for gene cloning. These techniques allowed the isolation of large amounts of a pure DNA fragment, free from all the other DNA sequences that together constitute the organism’s genome (all the genes in the chromosomes). This process enabled a DNA fragment—perhaps representing a particular gene—to be generated and characterized.

Gene cloning was coupled with the development of hybridization procedures in which a cloned DNA molecule is radioactively labeled and then made single-stranded. Some techniques use nonradioactive labels. The resulting molecule will bind by base pairing to any DNA or RNA that contains the same linear order of the four bases. As a result, it can be used as a probe to locate a particular DNA sequence within a DNA sample. The procedure used to accomplish this is known as Southern blotting after its inventor, Ed Southern.

In the related technique of Northern blotting, DNA from a gene is hybridized to the RNA prepared from different tissues, which allows the RNA corresponding to the gene to be detected and quantified in different tissues. These techniques have revealed much information on gene structure and expression.