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Plasmid Preparation
A plasmid preparation is a method of DNA extraction and purification for plasmid DNA. Many methods have been developed to purify plasmid DNA from bacteria that directly exist in the environment or bacteria that grown at laboratory level. These methods invariably involve three steps: * Growth of the bacterial culture * Harvesting and lysis of the bacteria * Purification of plasmid DNA Plasmid purification and preparation from the environment Plasmid purification directly from the environment is an important tool in molecular biology and biotechnology, and helps to advance our understanding of the role of plasmids in the biology of different organisms. In recent years, the use of cultivation-independent methods such as total community DNA (TC-DNA) extraction has allowed for a more comprehensive understanding of the prevalence and distribution of plasmids in the environment. This approach involves extracting DNA directly from environmental samples, allowing for the detection and ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Plasmid Miniprep
A plasmid is a small, extrachromosomal DNA molecule within a cell that is physically separated from chromosomal DNA and can replicate independently. They are most commonly found as small circular, double-stranded DNA molecules in bacteria; however, plasmids are sometimes present in archaea and eukaryotic organisms. In nature, plasmids often carry genes that benefit the survival of the organism and confer selective advantage such as antibiotic resistance. While chromosomes are large and contain all the essential genetic information for living under normal conditions, plasmids are usually very small and contain only additional genes that may be useful in certain situations or conditions. Artificial plasmids are widely used as vectors in molecular cloning, serving to drive the replication of recombinant DNA sequences within host organisms. In the laboratory, plasmids may be introduced into a cell via transformation. Synthetic plasmids are available for procurement over the inter ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Proteins
Proteins are large biomolecules and macromolecules that comprise one or more long chains of amino acid residues. Proteins perform a vast array of functions within organisms, including catalysing metabolic reactions, DNA replication, responding to stimuli, providing structure to cells and organisms, and transporting molecules from one location to another. Proteins differ from one another primarily in their sequence of amino acids, which is dictated by the nucleotide sequence of their genes, and which usually results in protein folding into a specific 3D structure that determines its activity. A linear chain of amino acid residues is called a polypeptide. A protein contains at least one long polypeptide. Short polypeptides, containing less than 20–30 residues, are rarely considered to be proteins and are commonly called peptides. The individual amino acid residues are bonded together by peptide bonds and adjacent amino acid residues. The sequence of amino acid residues ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Spin Column-based Nucleic Acid Purification
Spin column-based nucleic acid purification is a solid phase extraction method to quickly purify nucleic acids. This method relies on the fact that nucleic acid will bind to the solid phase of silica under certain conditions. Procedure The stages of the method are lyse, bind, wash, and elute. More specifically, this entails the lysis of target cells to release nucleic acids, selective binding of nucleic acid to a silica membrane, washing away particulates and inhibitors that are not bound to the silica membrane, and elution of the nucleic acid, with the end result being purified nucleic acid in an aqueous solution. For lysis, the cells (blood, tissue, etc.) of the sample must undergo a treatment to break the cell membrane and free the nucleic acid. Depending on the target material, this can include the use of detergent or other buffers, proteinases or other enzymes, heating to various times/temperatures, or mechanical disruption such as cutting with a knife or homogenizer, using ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Antisolvent
Salting out (also known as salt-induced precipitation, salt fractionation, anti-solvent crystallization, precipitation crystallization, or drowning out) is a purification technique that utilizes the reduced solubility of certain molecules in a solution of very high ionic strength. Salting out is typically used to precipitate large biomolecules, such as proteins or DNA. Because the salt concentration needed for a given protein to precipitate out of the solution differs from protein to protein, a specific salt concentration can be used to precipitate a target protein. This process is also used to concentrate dilute solutions of proteins. Dialysis can be used to remove the salt if needed. Principle Salt compounds dissociate in aqueous solutions. This property is exploited in the process of salting out. When the salt concentration is increased, some of the water molecules are attracted by the salt ions, which decreases the number of water molecules available to interact with the c ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Ethanol
Ethanol (abbr. EtOH; also called ethyl alcohol, grain alcohol, drinking alcohol, or simply alcohol) is an organic compound. It is an Alcohol (chemistry), alcohol with the chemical formula . Its formula can be also written as or (an ethyl group linked to a hydroxyl group). Ethanol is a Volatility (chemistry), volatile, Combustibility and flammability, flammable, colorless liquid with a characteristic wine-like odor and pungent taste. It is a psychoactive recreational drug, the active ingredient in alcoholic drinks. Ethanol is naturally produced by the fermentation process of Carbohydrate, sugars by yeasts or via Petrochemistry, petrochemical processes such as ethylene hydration. It has medical applications as an antiseptic and disinfectant. It is used as a chemical solvent and in the Chemical synthesis, synthesis of organic compounds, and as a Alcohol fuel, fuel source. Ethanol also can be dehydrated to make ethylene, an important chemical feedstock. As of 2006, world produ ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Ethanol Precipitation
Ethanol precipitation is a method used to purify and/or concentrate RNA, DNA, and polysaccharides such as pectin and xyloglucan from aqueous solutions by adding ethanol as an antisolvent. DNA precipitation Theory DNA is polar due to its highly charged phosphate backbone. Its polarity makes it water-soluble (water is polar) according to the principle "like dissolves like". Because of the high polarity of water, illustrated by its high dielectric constant of 80.1 (at 20 °C), electrostatic forces between charged particles are considerably lower in aqueous solution than they are in a vacuum or in air. This relation is reflected in Coulomb's law, which can be used to calculate the force acting on two charges q_1 and q_2 separated by a distance r by using the dielectric constant \varepsilon_r (also called relative static permittivity) of the medium in the denominator of the equation (\varepsilon_0 is an electric constant): F = \frac \frac At an atomic level, the reduction ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Biomolecule
A biomolecule or biological molecule is a loosely used term for molecules present in organisms that are essential to one or more typically biological processes, such as cell division, morphogenesis, or development. Biomolecules include large macromolecules (or polyelectrolytes) such as proteins, carbohydrates, lipids, and nucleic acids, as well as small molecules such as primary metabolites, secondary metabolites and natural products. A more general name for this class of material is biological materials. Biomolecules are an important element of living organisms, those biomolecules are often endogenous, produced within the organism but organisms usually need exogenous biomolecules, for example certain nutrients, to survive. Biology and its subfields of biochemistry and molecular biology study biomolecules and their reactions. Most biomolecules are organic compounds, and just four elements—oxygen, carbon, hydrogen, and nitrogen—make up 96% of the human body's mass. But ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Lysogeny Broth
Lysogeny broth (LB) is a nutritionally rich medium primarily used for the growth of bacteria. Its creator, Giuseppe Bertani, intended LB to stand for lysogeny broth, but LB has also come to colloquially mean Luria broth, Lennox broth, life broth or Luria–Bertani medium. The formula of the LB medium was published in 1951 in the first paper of Bertani on lysogeny. In this article he described the modified single-burst experiment and the isolation of the phages P1, P2, and P3. He had developed the LB medium to optimize ''Shigella'' growth and plaque formation. LB media formulations have been an industry standard for the cultivation of ''Escherichia coli'' as far back as the 1950s. These media have been widely used in molecular microbiology applications for the preparation of plasmid DNA and recombinant proteins. It continues to be one of the most common media used for maintaining and cultivating laboratory recombinant strains of ''Escherichia coli''. For physiological studi ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Cloning
Cloning is the process of producing individual organisms with identical or virtually identical DNA, either by natural or artificial means. In nature, some organisms produce clones through asexual reproduction. In the field of biotechnology, cloning is the process of creating cloned organisms (copies) of Cell (biology), cells and of DNA fragments (molecular cloning). Etymology Coined by Herbert J. Webber, the term clone derives from the Ancient Greek word (), ''twig'', which is the process whereby a new plant is created from a twig. In botany, the term ''lusus'' was used. In horticulture, the spelling ''clon'' was used until the early twentieth century; the final ''e'' came into use to indicate the vowel is a "long o" instead of a "short o". Since the term entered the popular lexicon in a more general context, the spelling ''clone'' has been used exclusively. Natural cloning Cloning is a natural form of reproduction that has allowed life forms to spread for hundreds of millio ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Molecular Cloning
Molecular cloning is a set of experimental methods in molecular biology that are used to assemble recombinant DNA molecules and to direct their replication within host organisms. The use of the word ''cloning'' refers to the fact that the method involves the replication of one molecule to produce a population of cells with identical DNA molecules. Molecular cloning generally uses DNA sequences from two different organisms: the species that is the source of the DNA to be cloned, and the species that will serve as the living host for replication of the recombinant DNA. Molecular cloning methods are central to many contemporary areas of modern biology and medicine. In a conventional molecular cloning experiment, the DNA to be cloned is obtained from an organism of interest, then treated with enzymes in the test tube to generate smaller DNA fragments. Subsequently, these fragments are then combined with vector DNA to generate recombinant DNA molecules. The recombinant DNA is then in ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Alkaline Lysis
Alkaline lysis or alkaline extraction is a method used in molecular biology to isolate plasmid DNA from bacteria. Method Bacteria containing the plasmid of interest are first cultured, then a sample is centrifuged in order to concentrate cellular material (including DNA) into a pellet at the bottom of the containing vessel. The supernatant is discarded, and the pellet is then re-suspended in an EDTA-containing physiological buffer. The purpose of the EDTA is to chelate divalent metal cations such as Mg2+ and Ca2+, which are required for the function of DNA degrading enzymes ( DNAses) and also serve to de-stabilise the DNA phosphate backbone and cell wall. Glucose in the buffer will maintain the osmotic pressure of the cell in order to prevent the cell from bursting. Tris in the buffer will retain the pH of the cell with 8.0 and RNase will remove the RNA which will disrupt the experiment. Separately, a strong alkaline solution consisting of the detergent sodium dodecyl sulfate ( ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Chromosomal
A chromosome is a long DNA molecule with part or all of the genetic material of an organism. In most chromosomes the very long thin DNA fibers are coated with packaging proteins; in eukaryotic cells the most important of these proteins are the histones. These proteins, aided by chaperone proteins, bind to and condense the DNA molecule to maintain its integrity. These chromosomes display a complex three-dimensional structure, which plays a significant role in transcriptional regulation. Chromosomes are normally visible under a light microscope only during the metaphase of cell division (where all chromosomes are aligned in the center of the cell in their condensed form). Before this happens, each chromosome is duplicated (S phase), and both copies are joined by a centromere, resulting either in an X-shaped structure (pictured above), if the centromere is located equatorially, or a two-arm structure, if the centromere is located distally. The joined copies are now called sis ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
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