Introduction to Methods in Molecular Biology

Today, the field of molecular biology focuses on the mechanisms by which cellular processes are carried out by the various biological macromolecules in the cell, with a particular emphasis on the structure and function of genes and genomes. Molecular biology as a field, however, was originally born from the development of tools and methods that allow the direct manipulation of DNA both in vitro and in vivo in numerous organisms.
Two essential items in the molecular biologist’s toolkit are restriction endonucleases, which allow DNA to be cut into precise pieces, and cloning vectors, such as plasmids or phages used to “carry” inserted foreign DNA fragments for the purpose of producing more material or a protein product. The term genetic engineering was originally used to describe the range of manipulations of DNA that become possible with the ability to clone a gene by placing its DNA into another context in which it could be propagated. From this beginning, when recombinant DNA was used  as a tool to analyze gene structure and expression, we moved to the ability to change the DNA content of bacteria and eukaryotic cells by directly introducing cloned DNA that could become part of the genome. Then, by changing the genetic content in conjunction with the ability to develop an animal from an embryonic cell, it became possible to generate multicellular eukaryotes with deletions or additions of specific genes that are inherited via the germline.

We now use genetic engineering to describe a range of activities  including the manipulation of DNA, the introduction of changes into specific somatic cells within an animal or plant, and even changes in the germline itself.
As research has advanced, more and more sensitive methods for detecting and amplifying DNA have been developed. Now that we have entered the era of routine whole-genome sequencing, the function and expression of entire genomes have become commonplace. This chapter discusses some of the most common methods used in molecular biology, ranging from the very first tools developed by molecular biologists to some of the most recently developed methods to assess the content.