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Immunoprecipitation
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. Types Individual protein immunoprecipitation (IP) Involves using an antibody that is specific for a known protein to isolate that particular protein out of a solution containing many different proteins. These solutions will often be in the form of a crude lysate of a plant or animal tissue. Other sample types could be body fluids or other samples of biological origin. Protein complex immunoprecipitation (Co-IP) Immunoprecipitation of intact protein complexes (i.e. antigen along with any proteins or ligands that are bound to it) is known as co-immunoprecipitation (Co-I ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Chromatin Immunoprecipitation
Chromatin immunoprecipitation (ChIP) is a type of immunoprecipitation experimental technique used to investigate the interaction between proteins and DNA in the cell. It aims to determine whether specific proteins are associated with specific genomic regions, such as transcription factors on promoters or other DNA binding sites, and possibly define cistromes. ChIP also aims to determine the specific location in the genome that various histone modifications are associated with, indicating the target of the histone modifiers. ChIP is crucial for the advancements in the field of epigenomics and learning more about epigenetic phenomena. Briefly, the conventional method is as follows: # DNA and associated proteins on chromatin in living cells or tissues are crosslinked (this step is omitted in Native ChIP). # The DNA-protein complexes (chromatin-protein) are then sheared into ~500 bp DNA fragments by sonication or nuclease digestion. # Cross-linked DNA fragments associated with the ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Antibody
An antibody (Ab), also known as an immunoglobulin (Ig), is a large, Y-shaped protein used by the immune system to identify and neutralize foreign objects such as pathogenic bacteria and Viral disease, viruses. The antibody recognizes a unique molecule of the pathogen, called an antigen. Each tip of the "Y" of an antibody contains a paratope (analogous to a lock) that is specific for one particular epitope (analogous to a key) on an antigen, allowing these two structures to bind together with precision. Using this binding mechanism, an antibody can ''tag'' a microbe or an infected cell for attack by other parts of the immune system, or can neutralize it directly (for example, by blocking a part of a virus that is essential for its invasion). To allow the immune system to recognize millions of different antigens, the antigen-binding sites at both tips of the antibody come in an equally wide variety. In contrast, the remainder of the antibody is relatively constant. It only occurs ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
PAR-CLIP
PAR-CLIP (photoactivatable ribonucleoside-enhanced crosslinking and immunoprecipitation) is a biochemical method for identifying the binding sites of cellular RNA-binding proteins (RBPs) and microRNA-containing ribonucleoprotein complexes (miRNPs). The method relies on the incorporation of ribonucleoside analogs that are photoreactive, such as 4-thiouridine (4-SU) and 6-thioguanosine (6-SG), into nascent RNA transcripts by living cells. Irradiation of the cells by ultraviolet light of 365 nm wavelength induces efficient crosslinking of photoreactive nucleoside– labeled cellular RNAs to interacting RBPs. Immunoprecipitation of the RBP of interest is followed by isolation of the crosslinked and coimmunoprecipitated RNA. The isolated RNA is converted into a cDNA library and is deep sequenced using next-generation sequencing technology. Recently, PAR-CLIP have been applied to determine the transcriptome-wide binding sites of several known RBPs and microRNA-containing ribonucle ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
ChIP-on-chip
ChIP-on-chip (also known as ChIP-chip) is a technology that combines chromatin immunoprecipitation ('ChIP') with DNA microarray (''"chip"''). Like regular ChIP, ChIP-on-chip is used to investigate interactions between proteins and DNA ''in vivo''. Specifically, it allows the identification of the cistrome, the sum of binding sites, for DNA-binding proteins on a genome-wide basis. Whole-genome analysis can be performed to determine the locations of binding sites for almost any protein of interest. As the name of the technique suggests, such proteins are generally those operating in the context of chromatin. The most prominent representatives of this class are transcription factors, replication-related proteins, like origin recognition complex protein (ORC), histones, their variants, and histone modifications. The goal of ChIP-on-chip is to locate protein binding sites that may help identify functional elements in the genome. For example, in the case of a transcription factor as ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Chromatin Immunoprecipitation Sequencing
Chromatin is a complex of DNA and protein found in eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser structures. This prevents the strands from becoming tangled and also plays important roles in reinforcing the DNA during cell division, preventing DNA damage, and regulating gene expression and DNA replication. During mitosis and meiosis, chromatin facilitates proper segregation of the chromosomes in anaphase; the characteristic shapes of chromosomes visible during this stage are the result of DNA being coiled into highly condensed chromatin. The primary protein components of chromatin are histones. An octamer of two sets of four histone cores ( Histone H2A, Histone H2B, Histone H3, and Histone H4) bind to DNA and function as "anchors" around which the strands are wound.Maeshima, K., Ide, S., & Babokhov, M. (2019). Dynamic chromatin organization without the 30-nm fiber. ''Current opinion in cell biology, 58,'' 95–104. https://do ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Precipitation (chemistry)
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 formatio ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Sonication
A sonicator at the Weizmann Institute of Science during sonicationSonication is the act of applying sound energy to agitate particles in a sample, for various purposes such as the extraction of multiple compounds from plants, microalgae and seaweeds. Ultrasonic frequencies (> 20 kHz) are usually used, leading to the process also being known as ultrasonication or ultra-sonication. In the laboratory, it is usually applied using an ''ultrasonic bath'' or an '' ultrasonic probe'', colloquially known as a ''sonicator''. In a paper machine, an ultrasonic foil can distribute cellulose fibres more uniformly and strengthen the paper. Effects Sonication has numerous effects, both chemical and physical. The chemical effects of ultrasound are concerned with understanding the effect of sonic waves on chemical systems, this is called sonochemistry. The chemical effects of ultrasound do not come from a direct interaction with molecular species. Studies have shown that no direct c ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Polymerase Chain Reaction
The polymerase chain reaction (PCR) is a method widely used to rapidly make millions to billions of copies (complete or partial) of a specific DNA sample, allowing scientists to take a very small sample of DNA and amplify it (or a part of it) to a large enough amount to study in detail. PCR was invented in 1983 by the American biochemist Kary Mullis at Cetus Corporation; Mullis and biochemist Michael Smith, who had developed other essential ways of manipulating DNA, were jointly awarded the Nobel Prize in Chemistry in 1993. PCR is fundamental to many of the procedures used in genetic testing and research, including analysis of ancient samples of DNA and identification of infectious agents. Using PCR, copies of very small amounts of DNA sequences are exponentially amplified in a series of cycles of temperature changes. PCR is now a common and often indispensable technique used in medical laboratory research for a broad variety of applications including biomedical research ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Plasmid
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 in ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Chip-Sequencing
ChIP-sequencing, also known as ChIP-seq, is a method used to analyze protein interactions with DNA. ChIP-seq combines chromatin immunoprecipitation (ChIP) with massively parallel DNA sequencing to identify the binding sites of DNA-associated proteins. It can be used to map global binding sites precisely for any protein of interest. Previously, ChIP-on-chip was the most common technique utilized to study these protein–DNA relations. Uses ChIP-seq is primarily used to determine how transcription factors and other chromatin-associated proteins influence phenotype-affecting mechanisms. Determining how proteins interact with DNA to regulate gene expression is essential for fully understanding many biological processes and disease states. This epigenetic information is complementary to genotype and expression analysis. ChIP-seq technology is currently seen primarily as an alternative to ChIP-chip which requires a hybridization array. This introduces some bias, as an array is restr ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
DNA Microarray
A DNA microarray (also commonly known as DNA chip or biochip) is a collection of microscopic DNA spots attached to a solid surface. Scientists use DNA microarrays to measure the expression levels of large numbers of genes simultaneously or to genotype multiple regions of a genome. Each DNA spot contains picomoles (10−12 moles) of a specific DNA sequence, known as '' probes'' (or ''reporters'' or '' oligos''). These can be a short section of a gene or other DNA element that are used to hybridize a cDNA or cRNA (also called anti-sense RNA) sample (called ''target'') under high-stringency conditions. Probe-target hybridization is usually detected and quantified by detection of fluorophore-, silver-, or chemiluminescence-labeled targets to determine relative abundance of nucleic acid sequences in the target. The original nucleic acid arrays were macro arrays approximately 9 cm × 12 cm and the first computerized image based analysis was published in 1981. It was i ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Cistrome
In simple words, the cistrome refers a collection of regulatory elements of a set of genes, including transcription factor binding-sites and histone modifications. More specifically, "the set of cis-acting targets of a trans-acting factor on a genome-wide scale, also known as the ''in vivo'' genome-wide location of transcription factor binding-sites or histone modifications". The term cistrome is a portmanteau of cistr (from cistron) + ome (from genome). The term cistrome was coined by investigators at the Dana–Farber Cancer Institute and Harvard Medical School. Technologies such as chromatin immunoprecipitation combined with microarray analysis "ChIP-on-chip" or with massively parallel DNA sequencing " ChIP-Seq" have greatly facilitated the definition of the cistrome of transcription factors and other chromatin Chromatin is a complex of DNA and protein found in eukaryote, eukaryotic cells. The primary function is to package long DNA molecules into more compact, denser struct ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
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