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DNA Separation By Silica Adsorption
DNA separation by silica adsorption is a method of DNA separation that is based on DNA molecules binding to silica surfaces in the presence of certain salts and under certain pH conditions. Operations In order to conduct DNA separation by silica adsorption, a sample (this may be anything from purified cells to a tissue specimen) is lysed, releasing proteins, DNA, phospholipids, etc. from the cells. The remaining tissue is discarded. The supernatant containing the DNA is then exposed to silica in a solution with high ionic strength. The highest DNA adsorption efficiencies occur in the presence of buffer solution with a pH at or below the pKa of the surface silanol groups. The mechanism behind DNA adsorption onto silica is not fully understood; one possible explanation involves reduction of the silica surface's negative charge due to the high ionic strength of the buffer. This decrease in surface charge leads to a decrease in the electrostatic repulsion between the negatively cha ...
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Silica
Silicon dioxide, also known as silica, is an oxide of silicon with the chemical formula , most commonly found in nature as quartz and in various living organisms. In many parts of the world, silica is the major constituent of sand. Silica is one of the most complex and most abundant families of materials, existing as a compound of several minerals and as a synthetic product. Notable examples include fused quartz, fumed silica, silica gel, opal and aerogels. It is used in structural materials, microelectronics (as an Insulator (electricity), electrical insulator), and as components in the food and pharmaceutical industries. Structure In the majority of silicates, the silicon atom shows tetrahedral coordination geometry, tetrahedral coordination, with four oxygen atoms surrounding a central Si atomsee 3-D Unit Cell. Thus, SiO2 forms 3-dimensional network solids in which each silicon atom is covalently bonded in a tetrahedral manner to 4 oxygen atoms. In contrast, CO2 is a linear ...
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Lysis
Lysis ( ) is the breaking down of the membrane of a cell, often by viral, enzymic, or osmotic (that is, "lytic" ) mechanisms that compromise its integrity. A fluid containing the contents of lysed cells is called a ''lysate''. In molecular biology, biochemistry, and cell biology laboratories, cell cultures may be subjected to lysis in the process of purifying their components, as in protein purification, DNA extraction, RNA extraction, or in purifying organelles. Many species of bacteria are subject to lysis by the enzyme lysozyme, found in animal saliva, egg white, and other secretions. Phage lytic enzymes (lysins) produced during bacteriophage infection are responsible for the ability of these viruses to lyse bacterial cells. Penicillin and related β-lactam antibiotics cause the death of bacteria through enzyme-mediated lysis that occurs after the drug causes the bacterium to form a defective cell wall. If the cell wall is completely lost and the penicillin was used ...
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Protein
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 residue ...
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Phospholipid
Phospholipids, are a class of lipids whose molecule has a hydrophilic "head" containing a phosphate group and two hydrophobic "tails" derived from fatty acids, joined by an alcohol residue (usually a glycerol molecule). Marine phospholipids typically have omega-3 fatty acids EPA and DHA integrated as part of the phospholipid molecule. The phosphate group can be modified with simple organic molecules such as choline, ethanolamine or serine. Phospholipids are a key component of all cell membranes. They can form lipid bilayers because of their amphiphilic characteristic. In eukaryotes, cell membranes also contain another class of lipid, sterol, interspersed among the phospholipids. The combination provides fluidity in two dimensions combined with mechanical strength against rupture. Purified phospholipids are produced commercially and have found applications in nanotechnology and materials science. The first phospholipid identified in 1847 as such in biological tissues was lecith ...
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Supernatant
In an aqueous solution, precipitation is the process of transforming a dissolved substance into an insoluble solid from a super-saturated solution. The solid formed is called the precipitate. In case of an inorganic chemical reaction leading to precipitation, the chemical reagent causing the solid to form is called the ''precipitant''. The clear liquid remaining above the precipitated or the centrifuged solid phase is also called the 'supernate' or 'supernatant'. The notion of precipitation can also be extended to other domains of chemistry (organic chemistry and biochemistry) and even be applied to the solid phases (''e.g.'', metallurgy and alloys) when solid impurities segregate from a solid phase. Supersaturation The precipitation of a compound may occur when its concentration exceeds its solubility. This can be due to temperature changes, solvent evaporation, or by mixing solvents. Precipitation occurs more rapidly from a strongly supersaturated solution. The formati ...
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Guanidinium Chloride
Guanidinium chloride or guanidine hydrochloride, usually abbreviated GdmCl and sometimes GdnHCl or GuHCl, is the hydrochloride salt of guanidine. Structure Guanidinium chloride crystallizes in orthorhombic space group ''Pbca''. The crystal structure consists of a network of guanidinium cations and chloride anions linked by N–H···Cl hydrogen bonds. Acidity Guanidinium chloride is a weak acid with a pKa of 13.6. The reason that it is such a weak acid is the complete delocalisation of the positive charge through 3 nitrogen atoms (plus a little bit positive charge on carbon). However, some stronger bases can deprotonate it, such as sodium hydroxide: C(NH2)3+ + OH- > HNC(NH2)2 + H2O The equilibrium is not complete because the acidity difference between guanidinium and water is not large (The approximate pKa values: 13.6 vs 15.7). Complete deprotonation should be done with extremely strong bases, such as lithium diisopropylamide. C(NH2)3+Cl- + Li+N(C3H7)2- -> HNC(NH2)2 + ...
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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 ...
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Biotechnology
Biotechnology is the integration of natural sciences and engineering sciences in order to achieve the application of organisms, cells, parts thereof and molecular analogues for products and services. The term ''biotechnology'' was first used by Károly Ereky in 1919, meaning the production of products from raw materials with the aid of living organisms. Definition The concept of biotechnology encompasses a wide range of procedures for modifying living organisms according to human purposes, going back to domestication of animals, cultivation of the plants, and "improvements" to these through breeding programs that employ artificial selection and hybridization. Modern usage also includes genetic engineering as well as cell and tissue culture technologies. The American Chemical Society defines biotechnology as the application of biological organisms, systems, or processes by various industries to learning about the science of life and the improvement of the value of materials ...
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Molecular Biology
Molecular biology is the branch of biology that seeks to understand the molecular basis of biological activity in and between cells, including biomolecular synthesis, modification, mechanisms, and interactions. The study of chemical and physical structure of biological macromolecules is known as molecular biology. Molecular biology was first described as an approach focused on the underpinnings of biological phenomena - uncovering the structures of biological molecules as well as their interactions, and how these interactions explain observations of classical biology. In 1945 the term molecular biology was used by physicist William Astbury. In 1953 Francis Crick, James Watson, Rosalind Franklin, and colleagues, working at Medical Research Council unit, Cavendish laboratory, Cambridge (now the MRC Laboratory of Molecular Biology), made a double helix model of DNA which changed the entire research scenario. They proposed the DNA structure based on previous research done by Ro ...
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