in modern biological engineering and industrial microbiology. The work of Stanley Norman Cohen and Herbert Boyer in E. coli, using plasmids and restriction enzymes to create recombinant DNA, became a foundation of biotechnology. Considered a very versatile host for the production of heterologous proteins, researchers can introduce genes into the microbes using plasmids, allowing for the mass production of proteins in industrial fermentation processes. Genetic systems have also been developed which allow the production of recombinant proteins using E. coli. One of the first useful applications of recombinant DNA technology was the manipulation of E. coli to produce human insulin. Modified E. coli have been used in vaccine development, bioremediation, and production of immobilised enzymes. E. coli have been used successfully to produce proteins previously thought difficult or impossible in E. coli, such as those containing multiple disulfide bonds or those requiring post-translational modification for stability or function. The cellular environment of E. coli is normally too reducing for disulphide bonds to form, proteins with disulphide bonds therefore may be secreted to its periplasmic space, however, mutants in which the reduction of both thioredoxins and glutathione is impaired also allow disulphide bonded proteins to be produced in the cytoplasm of E. coli. It has also been used to produce proteins with various post-translational modifications, including glycoproteins by using the N-linked glycosylation system of Campylobacter jejuni engineered into E. coli. Efforts are currently under way to expand this technology to produce complex glycosylations. Studies are also being performed into programming E. coli to potentially solve complicated mathematics problems such as the Hamiltonian path problem. Model organism E. coli is frequently used as a model organism in microbiology studies. Cultivated strains (e.g. E. coli K-12) are well-adapted to the laboratory environment, and, unlike wild type strains, have lost their ability to thrive in the intestine. Many lab strains lose their ability to form biofilms. These features protect wild type strains from antibodies and other chemical attacks, but require a large expenditure of energy and material resources. In 1946, Joshua Lederberg and Edward Tatum first described the phenomenon known as bacterial conjugation using E. coli as a model bacterium, and it remains a primary model to study conjugation. E. coli was an integral part of the first experiments to understand phage genetics, and early researchers, such as Seymour Benzer, used E. coli and phage T4 to understand the topography of gene structure. Prior