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Two-hybrid
Two-hybrid screening (originally known as yeast two-hybrid system or Y2H) is a molecular biology technique used to discover protein–protein interactions (PPIs) and protein–DNA interactions by testing for physical interactions (such as binding) between two proteins or a single protein and a DNA molecule, respectively. The premise behind the test is the activation of downstream reporter gene(s) by the binding of a transcription factor onto an upstream activating sequence (UAS). For two-hybrid screening, the transcription factor is split into two separate fragments, called the DNA-binding domain (DBD or often also abbreviated as BD) and activating domain (AD). The BD is the domain responsible for binding to the UAS and the AD is the domain responsible for the activation of transcription. The Y2H is thus a protein-fragment complementation assay. History Pioneered by Stanley Fields and Ok-Kyu Song in 1989, the technique was originally designed to detect protein–prote ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Protein–protein Interaction
Protein–protein interactions (PPIs) are physical contacts of high specificity established between two or more protein molecules as a result of biochemical events steered by interactions that include electrostatic forces, hydrogen bonding and the hydrophobic effect. Many are physical contacts with molecular associations between chains that occur in a cell or in a living organism in a specific biomolecular context. Proteins rarely act alone as their functions tend to be regulated. Many molecular processes within a cell are carried out by molecular machines that are built from numerous protein components organized by their PPIs. These physiological interactions make up the so-called interactomics of the organism, while aberrant PPIs are the basis of multiple aggregation-related diseases, such as Creutzfeldt–Jakob and Alzheimer's diseases. PPIs have been studied with many methods and from different perspectives: biochemistry, quantum chemistry, molecular dynamics, signal trans ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Protein-fragment Complementation Assay
Within the field of molecular biology, a protein-fragment complementation assay, or PCA, is a method for the identification and quantification of protein–protein interactions. In the PCA, the proteins of interest ("bait" and "prey") are each covalently linked to fragments of a third protein (e.g. DHFR, which acts as a "reporter"). Interaction between the bait and the prey proteins brings the fragments of the reporter protein in close proximity to allow them to form a functional reporter protein whose activity can be measured. This principle can be applied to many different reporter proteins and is also the basis for the yeast two-hybrid system, an archetypical PCA assay. Split protein assays Any protein that can be split into two parts and reconstituted non-covalently to form a functional protein may be used in a PCA. The two fragments however have low affinity for each other and must be brought together by other interacting proteins fused to them (often called "bait" and "pre ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
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 ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Reporter Gene
In molecular biology, a reporter gene (often simply reporter) is a gene that researchers attach to a regulatory sequence of another gene of interest in bacteria, cell culture, animals or plants. Such genes are called reporters because the characteristics they confer on organisms expressing them are easily identified and measured, or because they are selectable markers. Reporter genes are often used as an indication of whether a certain gene has been taken up by or expressed in the cell or organism population. Common reporter genes To introduce a reporter gene into an organism, scientists place the reporter gene and the gene of interest in the same DNA construct to be inserted into the cell or organism. For bacteria or prokaryotic cells in culture, this is usually in the form of a circular DNA molecule called a plasmid. For viruses, this is known as a viral vector. It is important to use a reporter gene that is not natively expressed in the cell or organism under study, since the ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Escherichia Coli
''Escherichia coli'' (),Wells, J. C. (2000) Longman Pronunciation Dictionary. Harlow ngland Pearson Education Ltd. also known as ''E. coli'' (), 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 ( EPEC, ETEC etc.) can cause serious food poisoning in their hosts, and are occasionally responsible for food contamination incidents that prompt product recalls. Most strains do not cause disease in humans and are part of the normal microbiota of the gut; such strains 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. col ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Saccharomyces Cerevisiae
''Saccharomyces cerevisiae'' () (brewer's yeast or baker's yeast) is a species of yeast (single-celled fungus microorganisms). The species has been instrumental in winemaking, baking, and brewing since ancient times. It is believed to have been originally isolated from the skin of grapes. It is one of the most intensively studied eukaryotic model organisms in molecular biology, molecular and cell biology, much like ''Escherichia coli'' as the model bacteria, bacterium. It is the microorganism behind the most common type of fermentation (biochemistry), fermentation. ''S. cerevisiae'' cells are round to ovoid, 5–10 micrometre, μm in diameter. It reproduces by budding. Many proteins important in human biology were first discovered by studying their Homology (biology), homologs in yeast; these proteins include cell cycle proteins, signaling proteins, and protein-processing enzymes. ''S. cerevisiae'' is currently the only yeast cell known to have Berkeley body, Berkeley bo ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
High-throughput Screening
High-throughput screening (HTS) is a method for scientific experimentation especially used in drug discovery and relevant to the fields of biology, materials science and chemistry. Using robotics, data processing/control software, liquid handling devices, and sensitive detectors, high-throughput screening allows a researcher to quickly conduct millions of chemical, genetic, or pharmacological tests. Through this process one can quickly recognize active compounds, antibodies, or genes that modulate a particular biomolecular pathway. The results of these experiments provide starting points for drug design and for understanding the noninteraction or role of a particular location. Assay plate preparation The key labware or testing vessel of HTS is the microtiter plate, which is a small container, usually disposable and made of plastic, that features a grid of small, open divots called ''wells''. In general, microplates for HTS have either 96, 192, 384, 1536, 3456 or 6144 wells. The ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Gal4
The Gal4 transcription factor is a positive regulator of gene expression of galactose-induced genes. This protein represents a large fungal family of transcription factors, Gal4 family, which includes over 50 members in the yeast ''Saccharomyces cerevisiae'' e.g. Oaf1, Pip2, Pdr1, Pdr3, Leu3. Gal4 recognizes genes with UAS, an upstream activating sequence, and activates them. In yeast cells, the principal targets are GAL1 (galactokinase), GAL10 (UDP-glucose 4-epimerase), and GAL7 (galactose-1-phosphate uridylyltransferase), three enzymes required for galactose metabolism. This binding has also proven useful in constructing the GAL4/UAS system, a technique for controlling expression in insects. In yeast, Gal4 is by default repressed by Gal80, and activated in the presence of galactose as Gal3 binds away Gal80. Domains Two executive domains, DNA binding and activation domains, provide key function of the Gal4 protein conforming to most of the transcription factors. DNA binding ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Genetic Engineering
Genetic engineering, also called genetic modification or genetic manipulation, is the modification and manipulation of an organism's genes using technology. It is a set of technologies used to change the genetic makeup of cells, including the transfer of genes within and across species boundaries to produce improved or novel organisms. New DNA is obtained by either isolating and copying the genetic material of interest using recombinant DNA methods or by artificially synthesising the DNA. A construct is usually created and used to insert this DNA into the host organism. The first recombinant DNA molecule was made by Paul Berg in 1972 by combining DNA from the monkey virus SV40 with the lambda virus. As well as inserting genes, the process can be used to remove, or "knock out", genes. The new DNA can be inserted randomly, or targeted to a specific part of the genome. An organism that is generated through genetic engineering is considered to be genetically modified (GM) an ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Biosynthesis
Biosynthesis is a multi-step, enzyme-catalyzed process where substrates are converted into more complex products in living organisms. In biosynthesis, simple compounds are modified, converted into other compounds, or joined to form macromolecules. This process often consists of metabolic pathways. Some of these biosynthetic pathways are located within a single cellular organelle, while others involve enzymes that are located within multiple cellular organelles. Examples of these biosynthetic pathways include the production of lipid membrane components and nucleotides. Biosynthesis is usually synonymous with anabolism. The prerequisite elements for biosynthesis include: precursor compounds, chemical energy (e.g. ATP), and catalytic enzymes which may require coenzymes (e.g.NADH, NADPH). These elements create monomers, the building blocks for macromolecules. Some important biological macromolecules include: proteins, which are composed of amino acid monomers joined via peptide bon ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Amino Acid
Amino acids are organic compounds that contain both amino and carboxylic acid functional groups. Although hundreds of amino acids exist in nature, by far the most important are the alpha-amino acids, which comprise proteins. Only 22 alpha amino acids appear in the genetic code. Amino acids can be classified according to the locations of the core structural functional groups, as Alpha and beta carbon, alpha- , beta- , gamma- or delta- amino acids; other categories relate to Chemical polarity, polarity, ionization, and side chain group type (aliphatic, Open-chain compound, acyclic, aromatic, containing hydroxyl or sulfur, etc.). In the form of proteins, amino acid '' residues'' form the second-largest component (water being the largest) of human muscles and other tissues. Beyond their role as residues in proteins, amino acids participate in a number of processes such as neurotransmitter transport and biosynthesis. It is thought that they played a key role in enabling life ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
Two Hybrid Assay
2 (two) is a number, numeral and digit. It is the natural number following 1 and preceding 3. It is the smallest and only even prime number. Because it forms the basis of a duality, it has religious and spiritual significance in many cultures. Evolution Arabic digit The digit used in the modern Western world to represent the number 2 traces its roots back to the Indic Brahmic script, where "2" was written as two horizontal lines. The modern Chinese and Japanese languages (and Korean Hanja) still use this method. The Gupta script rotated the two lines 45 degrees, making them diagonal. The top line was sometimes also shortened and had its bottom end curve towards the center of the bottom line. In the Nagari script, the top line was written more like a curve connecting to the bottom line. In the Arabic Ghubar writing, the bottom line was completely vertical, and the digit looked like a dotless closing question mark. Restoring the bottom line to its original horizontal ... [...More Info...] [...Related Items...] OR: [Wikipedia] [Google] [Baidu] |
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