During the last few decades, techniques for manipulating eukaryotic as well as prokaryotic DNA have witnessed a remarkable development. There are three main phases of the development of these techniques, which include recombinant DNA technology and gene cloning; polymerase chain reaction and DNA chips and microarrays. Using recombinant DNA technology, we can isolate and clone single copy of a gene or a DNA segment into an indefinite number of copies, all identical. This became possible, because bacteria, phages and plasmids reproduce in their usual style, even after insertion of foreign DNA, so that the inserted DNA also replicates faithfully with the parent DNA. This technique is called gene cloning. It involves the production of a large number of identical DNA molecules from a single ancestral DNA molecule. The essential characteristic of gene cloning is that the desired gene or DNA fragments must be selectively amplified resulting in a large increase in copy number of selected DNA sequences. In practice, this involves multiple rounds of DNA replication catalyzed by a DNA polymerase acting on one or more types of template DNA molecule.
In gene cloning experiments, the gene or any DNA segment is first inserted into a vector, but this chimaeric vector with foreign gene is incapable of multiplying, so that it is transferred into living cells of bacteria or eukaryotes, where it can multiply with the host cells. These host cells can be a variety of cells and the choice will usually depend upon the objective and limitation which a vector or a host cell may impose. Essentially, two different gene cloning approaches are used: cell-based gene cloning and cell free DNA cloning. Cell-based gene cloning is an in-vivo cloning method while cell-free gene cloning is an in vitro cloning method which involves polymerase gene reaction which involves enzyme mediated gene cloning. Utilizing these techniques, thousands of genes or DNA sequences from prokaryotes and eukaryotes including plants and animals have been cloned since 1970s. yhese cloned sequences, whether genes or regulatory sequences or random sequences have been used for a variety of purposes, both for basic studies and applied uses including commercial uses. They have primarily proved useful in the study of function of regulatory DNA sequences and the application of gene cloning in areas like medicine, agriculture and industry. In the industry, the cloned genes led to the production of useful drugs, which will compete in the commercial market, due to their production at a very low price. These strategies have been used in several cases and some of the cloned genes include cloning of nitrogen fixation (nif) genes; cloning of penicillin G acylase gene; cloning of Hepatitis B virus genome; cloning of genes for human peptide hormones (insulin and human growth hormone-HGH), etc. In some cases, the bacterial cells also transcribe the recombinant DNA molecules into mRNA, which is, in turn, translated into polypeptide chains. Hence, cloning can be employed not only to produce large quantities of the DNA sequence of an individual gene but also large amounts of the genes protein product as well.