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The first isolation of deoxyribonucleic acid (DNA) was done in 1869 by Friedrich Miescher. DNA extraction is the process of isolating DNA from the cells of an organism isolated from a sample, typically a biological sample such as blood, saliva, or tissue. It involves breaking open the cells, removing proteins and other contaminants, and purifying the DNA so that it is free of other cellular components. The purified DNA can then be used for downstream applications such as PCR, sequencing, or cloning. Currently, it is a routine procedure in molecular biology or forensic analyses.
Affinity chromatography is a method of separating a biomolecule from a mixture, based on a highly specific macromolecular binding interaction between the biomolecule and another substance. The specific type of binding interaction depends on the biomolecule of interest; antigen and antibody, enzyme and substrate, receptor and ligand, or protein and nucleic acid binding interactions are frequently exploited for isolation of various biomolecules. Affinity chromatography is useful for its high selectivity and resolution of separation, compared to other chromatographic methods.
Immunoprecipitation (IP) is the technique of precipitating a protein antigen out of solution using an antibody that specifically binds to that particular protein. This process can be used to isolate and concentrate a particular protein from a sample containing many thousands of different proteins. Immunoprecipitation requires that the antibody be coupled to a solid substrate at some point in the procedure.
Immunomagnetic separation (IMS) is a laboratory tool that can efficiently isolate cells out of body fluid or cultured cells. It can also be used as a method of quantifying the pathogenicity of food, blood or feces. DNA analysis have supported the combined use of both this technique and Polymerase Chain Reaction (PCR). Another laboratory separation tool is the affinity magnetic separation (AMS), which is more suitable for the isolation of prokaryotic cells.
Solid-phase extraction (SPE) is a solid-liquid extractive technique, by which compounds that are dissolved or suspended in a liquid mixture are separated, isolated or purified, from other compounds in this mixture, according to their physical and chemical properties. Analytical laboratories use solid phase extraction to concentrate and purify samples for analysis. Solid phase extraction can be used to isolate analytes of interest from a wide variety of matrices, including urine, blood, water, beverages, soil, and animal tissue.
Phenol extraction is a laboratory technique that purifies nucleic acid samples using a phenol solution. Phenol is common reagent in extraction because its properties allow for effective nucleic acid extraction, particularly as it strongly denatures proteins, it is a nucleic acid preservative, and it is immiscible in water.
A plasmid preparation is a method of DNA extraction and purification for plasmid DNA. It is an important step in many molecular biology experiments and is essential for the successful use of plasmids in research and biotechnology. Many methods have been developed to purify plasmid DNA from bacteria. During the purification procedure, the plasmid DNA is often separated from contaminating proteins and genomic DNA.
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 salt and ethanol as an antisolvent.
Acrydite is a phosphoramidite that allows the synthesis of oligonucleotides with a methacryl group at the 5' end. Acryl oligonucleotides have been tested, but the acrylyl group is not stable to storage. Acrydite-modified oligonucleotides can react with nucleophiles such as thiols, this forms the basis of the ez-rays chemistry which was used for microarrays. More importantly, Acrydite-modified oligonucleotides can be incorporated, stoichiometrically, into hydrogels such as polyacrylamide, using standard free radical polymerization chemistry, where the double bond in the Acrydite group reacts with other activated double bond containing compounds such as acrylamide.
Acid guanidinium thiocyanate-phenol-chloroform extraction is a liquid–liquid extraction technique in biochemistry and molecular biology. It is widely used for isolating RNA. This method may take longer than a column-based system such as the silica-based purification, but has higher purity and the advantage of high recovery of RNA. Furthermore, an RNA column is typically unsuitable for purification of short RNA species, such as siRNA, miRNA and tRNA.
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.
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.
Polony sequencing is an inexpensive but highly accurate multiplex sequencing technique that can be used to “read” millions of immobilized DNA sequences in parallel. This technique was first developed by Dr. George Church's group at Harvard Medical School. Unlike other sequencing techniques, Polony sequencing technology is an open platform with freely downloadable, open source software and protocols. Also, the hardware of this technique can be easily set up with a commonly available epifluorescence microscopy and a computer-controlled flowcell/fluidics system. Polony sequencing is generally performed on paired-end tags library that each molecule of DNA template is of 135 bp in length with two 17–18 bp paired genomic tags separated and flanked by common sequences. The current read length of this technique is 26 bases per amplicon and 13 bases per tag, leaving a gap of 4–5 bases in each tag.
The Juliá–Colonna epoxidation is an asymmetric poly-leucine catalyzed nucleophilic epoxidation of electron deficient olefins in a triphasic system. The reaction was reported by Sebastian Juliá at the Chemical Institute of Sarriá in 1980, with further elaboration by both Juliá and Stefano Colonna.
Massive parallel sequencing or massively parallel sequencing is any of several high-throughput approaches to DNA sequencing using the concept of massively parallel processing; it is also called next-generation sequencing (NGS) or second-generation sequencing. Some of these technologies emerged between 1993 and 1998 and have been commercially available since 2005. These technologies use miniaturized and parallelized platforms for sequencing of 1 million to 43 billion short reads per instrument run.
Illumina dye sequencing is a technique used to determine the series of base pairs in DNA, also known as DNA sequencing. The reversible terminated chemistry concept was invented by Bruno Canard and Simon Sarfati at the Pasteur Institute in Paris. It was developed by Shankar Balasubramanian and David Klenerman of Cambridge University, who subsequently founded Solexa, a company later acquired by Illumina. This sequencing method is based on reversible dye-terminators that enable the identification of single nucleotides as they are washed over DNA strands. It can also be used for whole-genome and region sequencing, transcriptome analysis, metagenomics, small RNA discovery, methylation profiling, and genome-wide protein-nucleic acid interaction analysis.
Thermoresponsive polymers can be used as stationary phase in liquid chromatography. Here, the polarity of the stationary phase can be varied by temperature changes, altering the power of separation without changing the column or solvent composition. Thermally related benefits of gas chromatography can now be applied to classes of compounds that are restricted to liquid chromatography due to their thermolability. In place of solvent gradient elution, thermoresponsive polymers allow the use of temperature gradients under purely aqueous isocratic conditions. The versatility of the system is controlled not only through changing temperature, but through the addition of modifying moieties that allow for a choice of enhanced hydrophobic interaction, or by introducing the prospect of electrostatic interaction. These developments have already introduced major improvements to the fields of hydrophobic interaction chromatography, size exclusion chromatography, ion exchange chromatography, and affinity chromatography separations as well as pseudo-solid phase extractions.
Boom method is a solid phase extraction method for isolating nucleic acid from a biological sample. This method is characterized by "absorbing the nucleic acids (NA) to the silica beads".
Microbeads, also called Ugelstad particles after the Norwegian chemist, professor John Ugelstad, who invented them in 1977 and patented the method in 1978, are uniform polymer particles, typically 0.5 to 500 microns in diameter. Bio-reactive molecules can be absorbed or coupled to their surface, and used to separate biological materials such as cells, proteins, or nucleic acids.
Diagnostic microbiology is the study of microbial identification. Since the discovery of the germ theory of disease, scientists have been finding ways to harvest specific organisms. Using methods such as differential media or genome sequencing, physicians and scientists can observe novel functions in organisms for more effective and accurate diagnosis of organisms. Methods used in diagnostic microbiology are often used to take advantage of a particular difference in organisms and attain information about what species it can be identified as, which is often through a reference of previous studies. New studies provide information that others can reference so that scientists can attain a basic understanding of the organism they are examining.
References
- 1 2 DeAngelis, M M; Wang, D G; Hawkins, T L (1995-11-25). "Solid-phase reversible immobilization for the isolation of PCR products". Nucleic Acids Research. 23 (22): 4742–4743. doi:10.1093/nar/23.22.4742. ISSN 0305-1048. PMC 307455 . PMID 8524672.
- ↑ Oberacker, Phil; Stepper, Peter; Bond, Donna M.; Höhn, Sven; Focken, Jule; Meyer, Vivien; Schelle, Luca; Sugrue, Victoria J.; Jeunen, Gert-Jan; Moser, Tim; Hore, Steven R. (2019-01-10). "Bio-On-Magnetic-Beads (BOMB): Open platform for high-throughput nucleic acid extraction and manipulation". PLOS Biology. 17 (1): e3000107. doi: 10.1371/journal.pbio.3000107 . ISSN 1544-9173. PMC 6343928 . PMID 30629605.