Escherichia coli in molecular biology

Last updated • 7 min readFrom Wikipedia, The Free Encyclopedia
E. coli colonies containing the fluorescent pGLO plasmid Fluorescent.jpg
E. coli colonies containing the fluorescent pGLO plasmid

Escherichia coli ( ˌɛʃɪˈrɪkiəˈkl ; commonly abbreviated E. coli) is a Gram-negative gammaproteobacterium commonly found in the lower intestine of warm-blooded organisms (endotherms). The descendants of two isolates, K-12 and B strain, are used routinely in molecular biology as both a tool and a model organism.

Contents

Diversity

Escherichia coli is one of the most diverse bacterial species, with several pathogenic strains with different symptoms and with only 20% of the genome common to all strains. [1] Furthermore, from the evolutionary point of view, the members of genus Shigella (dysenteriae, flexneri, boydii, sonnei) are actually E. coli strains "in disguise" (i.e. E. coli is paraphyletic to the genus). [2]

History

In 1885, Theodor Escherich, a German pediatrician, first discovered this species in the feces of healthy individuals and called it Bacterium coli commune because it is found in the colon and early classifications of Prokaryotes placed these in a handful of genera based on their shape and motility (at that time Ernst Haeckel's classification of Bacteria in the kingdom Monera was in place [3] ). [4]

Following a revision of Bacteria it was reclassified as Bacillus coli by Migula in 1895 [5] and later reclassified as Escherichia coli. [6]

Due to its ease of culture and fast doubling, it was used in the early microbiology experiments; however, bacteria were considered primitive and pre-cellular and received little attention before 1944, when Avery, Macleod and McCarty demonstrated that DNA was the genetic material using Salmonella typhimurium , following which Escherichia coli was used for linkage mapping studies. [7]

Strains

Four of the many E. coli strains (K-12, B, C, and W) are thought of as model organism strains. These are classified in Risk Group 1 in biosafety guidelines.[ citation needed ]

Escherich's isolate

The first isolate of Escherich was deposited in NCTC in 1920 by the Lister Institute in London (NCTC 86 Archived 2011-07-25 at the Wayback Machine ). [8]

K-12

A strain was isolated from a stool sample of a patient convalescent from diphtheria and was labelled K-12 (not an antigen) in 1922 at Stanford University. [9] This isolate was used in 1940s by Charles E. Clifton to study nitrogen metabolism, who deposited it in ATCC (strain ATCC 10798 Archived 2011-07-25 at the Wayback Machine ) and lent it to Edward Tatum for his tryptophan biosynthesis experiments, [10] despite its idiosyncrasies due to the F+ λ+ phenotype. [7] In the course of the passages it lost its O antigen [7] and in 1953 was cured first of its lambda phage (strain W1485 Archived 2011-07-25 at the Wayback Machine by UV by Joshua Lederberg and colleagues) and then in 1985 of the F plasmid by acridine orange curing.[ citation needed ] Strains derived from MG1655 include DH1, parent of DH5α and in turn of DH10B (rebranded as TOP10 by Invitrogen [11] ). [12] An alternative lineage from W1485 is that of W2637 (which contains an inversion rrnD-rrnE), which in turn resulted in W3110. [8] Due to the lack of specific record-keeping, the "pedigree" of strains was not available and had to be inferred by consulting lab-book and records in order to set up the E. coli Genetic Stock Centre at Yale by Barbara Bachmann. [9] The different strains have been derived through treating E. coli K-12 with agents such as nitrogen mustard, ultra-violet radiation, X-ray etc. An extensive list of Escherichia coli K-12 strain derivatives and their individual construction, genotypes, phenotypes, plasmids and phage information can be viewed at Ecoliwiki.

B strain

A second common laboratory strain is the B strain, whose history is less straightforward and the first naming of the strain as E. coli B was by Delbrück and Luria in 1942 in their study of bacteriophages T1 and T7. [13] The original E. coli B strain, known then as Bacillus coli, originated from Félix d'Herelle from the Institut Pasteur in Paris around 1918 who studied bacteriophages, [14] who claimed that it originated from Collection of the Institut Pasteur, [15] but no strains of that period exist. [8] The strain of d'Herelle was passed to Jules Bordet, Director of the Institut Pasteur du Brabant in Bruxelles [16] and his student André Gratia. [17] The former passed the strain to Ann Kuttner ("the Bact. coli obtained from Dr. Bordet") [18] and in turn to Eugène Wollman (B. coli Bordet), [19] whose son deposited it in 1963 (CIP 63.70) as "strain BAM" (B American), while André Gratia passed the strain to Martha Wollstein, a researcher at Rockefeller, who refers to the strain as "Brussels strain of Bacillus coli" in 1921, [20] who in turn passed it to Jacques Bronfenbrenner (B. coli P.C.), who passed it to Delbrück and Luria. [8] [13] This strain gave rise to several other strains, such as REL606 and BL21. [8]

C strain

E. coli C is morphologically distinct from other E. coli strains; it is more spherical in shape and has a distinct distribution of its nucleoid. [21]

W strain

The W strain was isolated from the soil near Rutgers University by Selman Waksman. [22]

Role in biotechnology

Because of its long history of laboratory culture and ease of manipulation, E. coli also plays an important role in modern biological engineering and industrial microbiology. [23] 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. [24]

Considered a very versatile host for the production of heterologous proteins, [25] 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. [26] Modified E. coli have been used in vaccine development, bioremediation, and production of immobilised enzymes. [25]

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. [27] 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. [28] [29] Efforts are currently under way to expand this technology to produce complex glycosylations. [30] [31]

Studies are also being performed into programming E. coli to potentially solve complicated mathematics problems such as the Hamiltonian path problem. [32]

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. [33] [34] These features protect wild type strains from antibodies and other chemical attacks, but require a large expenditure of energy and material resources.[ citation needed ]

In 1946, Joshua Lederberg and Edward Tatum first described the phenomenon known as bacterial conjugation using E. coli as a model bacterium, [35] and it remains a primary model to study conjugation. [36] E. coli was an integral part of the first experiments to understand phage genetics, [37] and early researchers, such as Seymour Benzer, used E. coli and phage T4 to understand the topography of gene structure. [38] Prior to Benzer's research, it was not known whether the gene was a linear structure, or if it had a branching pattern.[ citation needed ]

E. coli was one of the first organisms to have its genome sequenced; the complete genome of E. coli K-12 was published by Science in 1997. [39]

Lenski's long-term evolution experiment

The long-term evolution experiments using E. coli, begun by Richard Lenski in 1988, have allowed direct observation of major evolutionary shifts in the laboratory. [40] In this experiment, one population of E. coli unexpectedly evolved the ability to aerobically metabolize citrate. This capacity is extremely rare in E. coli. As the inability to grow aerobically is normally used as a diagnostic criterion with which to differentiate E. coli from other, closely related bacteria such as Salmonella , this innovation may mark a speciation event observed in the lab.[ citation needed ]

Related Research Articles

<span class="mw-page-title-main">Bacteriophage</span> Virus that infects and replicates within bacteria

A bacteriophage, also known informally as a phage, is a virus that infects and replicates within bacteria and archaea. The term was derived from "bacteria" and the Greek φαγεῖν, meaning "to devour". Bacteriophages are composed of proteins that encapsulate a DNA or RNA genome, and may have structures that are either simple or elaborate. Their genomes may encode as few as four genes and as many as hundreds of genes. Phages replicate within the bacterium following the injection of their genome into its cytoplasm.

<span class="mw-page-title-main">Bacterial conjugation</span> Method of bacterial gene transfer

Bacterial conjugation is the transfer of genetic material between bacterial cells by direct cell-to-cell contact or by a bridge-like connection between two cells. This takes place through a pilus. It is a parasexual mode of reproduction in bacteria.

<span class="mw-page-title-main">Lambda phage</span> Bacteriophage that infects Escherichia coli

Enterobacteria phage λ is a bacterial virus, or bacteriophage, that infects the bacterial species Escherichia coli. It was discovered by Esther Lederberg in 1950. The wild type of this virus has a temperate life cycle that allows it to either reside within the genome of its host through lysogeny or enter into a lytic phase, during which it kills and lyses the cell to produce offspring. Lambda strains, mutated at specific sites, are unable to lysogenize cells; instead, they grow and enter the lytic cycle after superinfecting an already lysogenized cell.

<i>Escherichia coli</i> Enteric, rod-shaped, gram-negative bacterium

Escherichia coli ( ESH-ə-RIK-ee-ə KOH-lye) is a gram-negative, facultative anaerobic, rod-shaped, coliform bacterium of the genus Escherichia that is commonly found in the lower intestine of warm-blooded organisms. Most E. coli strains are harmless, but some serotypes such as EPEC, and ETEC are pathogenic and can cause serious food poisoning in their hosts, and are occasionally responsible for food contamination incidents that prompt product recalls. Most strains are part of the normal microbiota of the gut and are harmless or even beneficial to humans (although these strains tend to be less studied than the pathogenic ones). For example, some strains of E. coli benefit their hosts by producing vitamin K2 or by preventing the colonization of the intestine by pathogenic bacteria. These mutually beneficial relationships between E. coli and humans are a type of mutualistic biological relationship — where both the humans and the E. coli are benefitting each other. E. coli is expelled into the environment within fecal matter. The bacterium grows massively in fresh fecal matter under aerobic conditions for three days, but its numbers decline slowly afterwards.

<span class="mw-page-title-main">Genetic transformation</span> Genetic alteration of a cell by uptake of genetic material from the environment

In molecular biology and genetics, transformation is the genetic alteration of a cell resulting from the direct uptake and incorporation of exogenous genetic material from its surroundings through the cell membrane(s). For transformation to take place, the recipient bacterium must be in a state of competence, which might occur in nature as a time-limited response to environmental conditions such as starvation and cell density, and may also be induced in a laboratory.

<span class="mw-page-title-main">Transduction (genetics)</span> Transfer process in genetics

Transduction is the process by which foreign DNA is introduced into a cell by a virus or viral vector. An example is the viral transfer of DNA from one bacterium to another and hence an example of horizontal gene transfer. Transduction does not require physical contact between the cell donating the DNA and the cell receiving the DNA, and it is DNase resistant. Transduction is a common tool used by molecular biologists to stably introduce a foreign gene into a host cell's genome.

<i>Escherichia virus T4</i> Species of bacteriophage

Escherichia virus T4 is a species of bacteriophages that infect Escherichia coli bacteria. It is a double-stranded DNA virus in the subfamily Tevenvirinae of the family Straboviridae. T4 is capable of undergoing only a lytic life cycle and not the lysogenic life cycle. The species was formerly named T-even bacteriophage, a name which also encompasses, among other strains, Enterobacteria phage T2, Enterobacteria phage T4 and Enterobacteria phage T6.

DNA gyrase, or simply gyrase, is an enzyme within the class of topoisomerase and is a subclass of Type II topoisomerases that reduces topological strain in an ATP dependent manner while double-stranded DNA is being unwound by elongating RNA-polymerase or by helicase in front of the progressing replication fork. It is the only known enzyme to actively contribute negative supercoiling to DNA, while it also is capable of relaxing positive supercoils. It does so by looping the template to form a crossing, then cutting one of the double helices and passing the other through it before releasing the break, changing the linking number by two in each enzymatic step. This process occurs in bacteria, whose single circular DNA is cut by DNA gyrase and the two ends are then twisted around each other to form supercoils. Gyrase is also found in eukaryotic plastids: it has been found in the apicoplast of the malarial parasite Plasmodium falciparum and in chloroplasts of several plants. Bacterial DNA gyrase is the target of many antibiotics, including nalidixic acid, novobiocin, albicidin, and ciprofloxacin.

<span class="mw-page-title-main">Phi X 174</span> A single-stranded DNA virus that infects bacteria

The phi X 174 bacteriophage is a single-stranded DNA (ssDNA) virus that infects Escherichia coli. This virus was isolated in 1935 by Nicolas Bulgakov in Félix d'Hérelle's laboratory at the Pasteur Institute, from samples collected in Paris sewers. Its characterization and the study of its replication mechanism were carried out from the 1950s onwards. It was the first DNA-based genome to be sequenced. This work was completed by Fred Sanger and his team in 1977. In 1962, Walter Fiers and Robert Sinsheimer had already demonstrated the physical, covalently closed circularity of ΦX174 DNA. Nobel prize winner Arthur Kornberg used ΦX174 as a model to first prove that DNA synthesized in a test tube by purified enzymes could produce all the features of a natural virus, ushering in the age of synthetic biology. In 1972–1974, Jerard Hurwitz, Sue Wickner, and Reed Wickner with collaborators identified the genes required to produce the enzymes to catalyze conversion of the single stranded form of the virus to the double stranded replicative form. In 2003, it was reported by Craig Venter's group that the genome of ΦX174 was the first to be completely assembled in vitro from synthesized oligonucleotides. The ΦX174 virus particle has also been successfully assembled in vitro. In 2012, it was shown how its highly overlapping genome can be fully decompressed and still remain functional.

<span class="mw-page-title-main">T7 phage</span> Species of virus

Bacteriophage T7 is a bacteriophage, a virus that infects bacteria. It infects most strains of Escherichia coli and relies on these hosts to propagate. Bacteriophage T7 has a lytic life cycle, meaning that it destroys the cell it infects. It also possesses several properties that make it an ideal phage for experimentation: its purification and concentration have produced consistent values in chemical analyses; it can be rendered noninfectious by exposure to UV light; and it can be used in phage display to clone RNA binding proteins.

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<span class="mw-page-title-main">Esther Lederberg</span> American microbiologist (1922–2006)

Esther Miriam Zimmer Lederberg was an American microbiologist and a pioneer of bacterial genetics. She discovered the bacterial virus lambda phage and the bacterial fertility factor F, devised the first implementation of replica plating, and furthered the understanding of the transfer of genes between bacteria by specialized transduction.

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A P1-derived artificial chromosome, or PAC, is a DNA construct derived from the DNA of P1 bacteriophages and Bacterial artificial chromosome. It can carry large amounts of other sequences for a variety of bioengineering purposes in bacteria. It is one type of the efficient cloning vector used to clone DNA fragments in Escherichia coli cells.

<span class="mw-page-title-main">Toxin-antitoxin system</span> Biological process

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<span class="mw-page-title-main">Daisy Roulland-Dussoix</span> Swiss microbiologist and molecular biologist (1936–2014)

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<span class="mw-page-title-main">Integration host factor</span>

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