International Standard Serial Number
An International Standard Serial Number is an eight-digit serial number used to uniquely identify a serial publication, such as a magazine. The ISSN is helpful in distinguishing between serials with the same title. ISSN are used in ordering, interlibrary loans, other practices in connection with serial literature; the ISSN system was first drafted as an International Organization for Standardization international standard in 1971 and published as ISO 3297 in 1975. ISO subcommittee TC 46/SC 9 is responsible for maintaining the standard; when a serial with the same content is published in more than one media type, a different ISSN is assigned to each media type. For example, many serials are published both in electronic media; the ISSN system refers to these types as electronic ISSN, respectively. Conversely, as defined in ISO 3297:2007, every serial in the ISSN system is assigned a linking ISSN the same as the ISSN assigned to the serial in its first published medium, which links together all ISSNs assigned to the serial in every medium.
The format of the ISSN is an eight digit code, divided by a hyphen into two four-digit numbers. As an integer number, it can be represented by the first seven digits; the last code digit, which may be 0-9 or an X, is a check digit. Formally, the general form of the ISSN code can be expressed as follows: NNNN-NNNC where N is in the set, a digit character, C is in; the ISSN of the journal Hearing Research, for example, is 0378-5955, where the final 5 is the check digit, C=5. To calculate the check digit, the following algorithm may be used: Calculate the sum of the first seven digits of the ISSN multiplied by its position in the number, counting from the right—that is, 8, 7, 6, 5, 4, 3, 2, respectively: 0 ⋅ 8 + 3 ⋅ 7 + 7 ⋅ 6 + 8 ⋅ 5 + 5 ⋅ 4 + 9 ⋅ 3 + 5 ⋅ 2 = 0 + 21 + 42 + 40 + 20 + 27 + 10 = 160 The modulus 11 of this sum is calculated. For calculations, an upper case X in the check digit position indicates a check digit of 10. To confirm the check digit, calculate the sum of all eight digits of the ISSN multiplied by its position in the number, counting from the right.
The modulus 11 of the sum must be 0. There is an online ISSN checker. ISSN codes are assigned by a network of ISSN National Centres located at national libraries and coordinated by the ISSN International Centre based in Paris; the International Centre is an intergovernmental organization created in 1974 through an agreement between UNESCO and the French government. The International Centre maintains a database of all ISSNs assigned worldwide, the ISDS Register otherwise known as the ISSN Register. At the end of 2016, the ISSN Register contained records for 1,943,572 items. ISSN and ISBN codes are similar in concept. An ISBN might be assigned for particular issues of a serial, in addition to the ISSN code for the serial as a whole. An ISSN, unlike the ISBN code, is an anonymous identifier associated with a serial title, containing no information as to the publisher or its location. For this reason a new ISSN is assigned to a serial each time it undergoes a major title change. Since the ISSN applies to an entire serial a new identifier, the Serial Item and Contribution Identifier, was built on top of it to allow references to specific volumes, articles, or other identifiable components.
Separate ISSNs are needed for serials in different media. Thus, the print and electronic media versions of a serial need separate ISSNs. A CD-ROM version and a web version of a serial require different ISSNs since two different media are involved. However, the same ISSN can be used for different file formats of the same online serial; this "media-oriented identification" of serials made sense in the 1970s. In the 1990s and onward, with personal computers, better screens, the Web, it makes sense to consider only content, independent of media; this "content-oriented identification" of serials was a repressed demand during a decade, but no ISSN update or initiative occurred. A natural extension for ISSN, the unique-identification of the articles in the serials, was the main demand application. An alternative serials' contents model arrived with the indecs Content Model and its application, the digital object identifier, as ISSN-independent initiative, consolidated in the 2000s. Only in 2007, ISSN-L was defined in the
Nucleotides are organic molecules that serve as the monomer units for forming the nucleic acid polymers deoxyribonucleic acid and ribonucleic acid, both of which are essential biomolecules within all life-forms on Earth. Nucleotides are the building blocks of nucleic acids. A nucleoside is a 5-carbon sugar, thus a nucleoside plus a phosphate group yields a nucleotide. Nucleotides play a central role in metabolism at a fundamental, cellular level, they carry packets of chemical energy—in the form of the nucleoside triphosphates Adenosine triphosphate, Guanosine triphosphate, Cytidine triphosphate and Uridine triphosphate —throughout the cell to the many cellular functions that demand energy, which include: synthesizing amino acids and cell membranes and parts, moving the cell and moving cell parts, dividing the cell, etc. In addition, nucleotides participate in cell signaling, are incorporated into important cofactors of enzymatic reactions. In experimental biochemistry, nucleotides can be radiolabeled with radionuclides to yield radionucleotides.
A nucleotide is composed of three distinctive chemical sub-units: a five-carbon sugar molecule, a nitrogenous base—which two together are called a nucleoside—and one phosphate group. With all three joined, a nucleotide is termed a "nucleoside monophosphate"; the chemistry sources ACS Style Guide and IUPAC Gold Book prescribe that a nucleotide should contain only one phosphate group, but common usage in molecular biology textbooks extends the definition to include molecules with two, or with three, phosphates. Thus, the terms "nucleoside diphosphate" or "nucleoside triphosphate" may indicate nucleotides. Nucleotides contain either a purine or a pyrimidine base—i.e. The nitrogenous base molecule known as a nucleobase—and are termed ribonucleotides if the sugar is ribose, or deoxyribonucleotides if the sugar is deoxyribose. Individual phosphate molecules repetitively connect the sugar-ring molecules in two adjacent nucleotide monomers, thereby connecting the nucleotide monomers of a nucleic acid end-to-end into a long chain.
These chain-joins of sugar and phosphate molecules create a'backbone' strand for a single- or double helix. In any one strand, the chemical orientation of the chain-joins runs from the 5'-end to the 3'-end —referring to the five carbon sites on sugar molecules in adjacent nucleotides. In a double helix, the two strands are oriented in opposite directions, which permits base pairing and complementarity between the base-pairs, all, essential for replicating or transcribing the encoded information found in DNA. Unlike in nucleic acid nucleotides, singular cyclic nucleotides are formed when the phosphate group is bound twice to the same sugar molecule, i.e. at the corners of the sugar hydroxyl groups. These individual nucleotides function in cell metabolism rather than the nucleic acid structures of long-chain molecules. Nucleic acids are polymeric macromolecules assembled from nucleotides, the monomer-units of nucleic acids; the purine bases adenine and guanine and pyrimidine base cytosine occur in both DNA and RNA, while the pyrimidine bases thymine and uracil in just one.
Adenine forms a base pair with thymine with two hydrogen bonds, while guanine pairs with cytosine with three hydrogen bonds. Nucleotides can be synthesized by a variety of means both in vitro and in vivo. In vitro, protecting groups may be used during laboratory production of nucleotides. A purified nucleoside is protected to create a phosphoramidite, which can be used to obtain analogues not found in nature and/or to synthesize an oligonucleotide. In vivo, nucleotides can be recycled through salvage pathways; the components used in de novo nucleotide synthesis are derived from biosynthetic precursors of carbohydrate and amino acid metabolism, from ammonia and carbon dioxide. The liver is the major organ of de novo synthesis of all four nucleotides. De novo synthesis of pyrimidines and purines follows two different pathways. Pyrimidines are synthesized first from aspartate and carbamoyl-phosphate in the cytoplasm to the common precursor ring structure orotic acid, onto which a phosphorylated ribosyl unit is covalently linked.
Purines, are first synthesized from the sugar template onto which the ring synthesis occurs. For reference, the syntheses of the purine and pyrimidine nucleotides are carried out by several enzymes in the cytoplasm of the cell, not within a specific organelle. Nucleotides undergo breakdown such that useful parts can be reused in synthesis reactions to create new nucleotides; the synthesis of the pyrimidines CTP and UTP occurs in the cytoplasm and starts with the formation of carbamoyl phosphate from glutamine and CO2. Next, aspartate carbamoyltransferase catalyzes a condensation reaction between aspartate and carbamoyl phosphate to form carbamoyl aspartic acid, cyclized into 4,5-dihydroorotic acid by dihydroorotase; the latter is converted to orotate by dihydroorotate oxidase. The net reaction is: -Dihydroorotate + O2 → Orotate + H2O2Orotate is covalently linked with a phosphorylated ribosyl unit; the covalent linkage between the ribose and pyrimidine occurs at position C1 of the ribose unit, which contains a pyrophosphate, N1 of the pyrimidine ring.
Orotate phosphoribosyltransferase catalyzes the net reaction yielding orotidine monophosphate: Or
Enzymes are macromolecular biological catalysts. Enzymes accelerate chemical reactions; the molecules upon which enzymes may act are called substrates and the enzyme converts the substrates into different molecules known as products. All metabolic processes in the cell need enzyme catalysis in order to occur at rates fast enough to sustain life. Metabolic pathways depend upon enzymes to catalyze individual steps; the study of enzymes is called enzymology and a new field of pseudoenzyme analysis has grown up, recognising that during evolution, some enzymes have lost the ability to carry out biological catalysis, reflected in their amino acid sequences and unusual'pseudocatalytic' properties. Enzymes are known to catalyze more than 5,000 biochemical reaction types. Most enzymes are proteins; the latter are called ribozymes. Enzymes' specificity comes from their unique three-dimensional structures. Like all catalysts, enzymes increase the reaction rate by lowering its activation energy; some enzymes can make their conversion of substrate to product occur many millions of times faster.
An extreme example is orotidine 5'-phosphate decarboxylase, which allows a reaction that would otherwise take millions of years to occur in milliseconds. Chemically, enzymes are like any catalyst and are not consumed in chemical reactions, nor do they alter the equilibrium of a reaction. Enzymes differ from most other catalysts by being much more specific. Enzyme activity can be affected by other molecules: inhibitors are molecules that decrease enzyme activity, activators are molecules that increase activity. Many therapeutic drugs and poisons are enzyme inhibitors. An enzyme's activity decreases markedly outside its optimal temperature and pH, many enzymes are denatured when exposed to excessive heat, losing their structure and catalytic properties; some enzymes are used commercially, in the synthesis of antibiotics. Some household products use enzymes to speed up chemical reactions: enzymes in biological washing powders break down protein, starch or fat stains on clothes, enzymes in meat tenderizer break down proteins into smaller molecules, making the meat easier to chew.
By the late 17th and early 18th centuries, the digestion of meat by stomach secretions and the conversion of starch to sugars by plant extracts and saliva were known but the mechanisms by which these occurred had not been identified. French chemist Anselme Payen was the first to discover an enzyme, diastase, in 1833. A few decades when studying the fermentation of sugar to alcohol by yeast, Louis Pasteur concluded that this fermentation was caused by a vital force contained within the yeast cells called "ferments", which were thought to function only within living organisms, he wrote that "alcoholic fermentation is an act correlated with the life and organization of the yeast cells, not with the death or putrefaction of the cells."In 1877, German physiologist Wilhelm Kühne first used the term enzyme, which comes from Greek ἔνζυμον, "leavened" or "in yeast", to describe this process. The word enzyme was used to refer to nonliving substances such as pepsin, the word ferment was used to refer to chemical activity produced by living organisms.
Eduard Buchner submitted his first paper on the study of yeast extracts in 1897. In a series of experiments at the University of Berlin, he found that sugar was fermented by yeast extracts when there were no living yeast cells in the mixture, he named the enzyme that brought about the fermentation of sucrose "zymase". In 1907, he received the Nobel Prize in Chemistry for "his discovery of cell-free fermentation". Following Buchner's example, enzymes are named according to the reaction they carry out: the suffix -ase is combined with the name of the substrate or to the type of reaction; the biochemical identity of enzymes was still unknown in the early 1900s. Many scientists observed that enzymatic activity was associated with proteins, but others argued that proteins were carriers for the true enzymes and that proteins per se were incapable of catalysis. In 1926, James B. Sumner crystallized it; the conclusion that pure proteins can be enzymes was definitively demonstrated by John Howard Northrop and Wendell Meredith Stanley, who worked on the digestive enzymes pepsin and chymotrypsin.
These three scientists were awarded the 1946 Nobel Prize in Chemistry. The discovery that enzymes could be crystallized allowed their structures to be solved by x-ray crystallography; this was first done for lysozyme, an enzyme found in tears and egg whites that digests the coating of some bacteria. This high-resolution structure of lysozyme marked the beginning of the field of structural biology and the effort to understand how enzymes work at an atomic level of detail. An enzyme's name is derived from its substrate or the chemical reaction it catalyzes, with the word ending in -ase. Examples are alcohol dehydrogenase and DNA polymerase. Different enzymes that catalyze the same chemical reaction are called isozymes; the International Union of Biochemistry and Molecular Biology have developed a nomenclature for enzymes, the EC numbers. The first number broadly classifies the enzyme based on its mechanism; the top-level classification is: EC 1, Oxidoreductases: catalyze oxidation/reducti
Nicotinamide adenine dinucleotide
Nicotinamide adenine dinucleotide is a cofactor found in all living cells. The compound is called a dinucleotide because it consists of two nucleotides joined through their phosphate groups. One nucleotide contains the other nicotinamide. Nicotinamide adenine dinucleotide exists in two forms: an oxidized and reduced form, abbreviated as NAD+ and NADH respectively. In metabolism, nicotinamide adenine dinucleotide is involved in redox reactions, carrying electrons from one reaction to another; the cofactor is, found in two forms in cells: NAD+ is an oxidizing agent – it accepts electrons from other molecules and becomes reduced. This reaction forms NADH, which can be used as a reducing agent to donate electrons; these electron transfer reactions are the main function of NAD. However, it is used in other cellular processes, most notably a substrate of enzymes that add or remove chemical groups from proteins, in posttranslational modifications; because of the importance of these functions, the enzymes involved in NAD metabolism are targets for drug discovery.
In organisms, NAD can be synthesized from simple building-blocks from the amino acids tryptophan or aspartic acid. In an alternative fashion, more complex components of the coenzymes are taken up from food as niacin. Similar compounds are released by reactions that break down the structure of NAD; these preformed components pass through a salvage pathway that recycles them back into the active form. Some NAD is converted into the coenzyme nicotinamide adenine dinucleotide phosphate; the chemistry of NADP is similar to that of NAD, but it has different role, being predominantly a cofactor in anabolic metabolism. NAD+ is written with a superscript plus sign because of the formal charge on one of its nitrogen atoms. NADH, on the other hand, is a doubly charged anion because of its two bridging phosphate groups. Nicotinamide adenine dinucleotide, like all dinucleotides, consists of two nucleosides joined by a pair of bridging phosphate groups; the nucleosides each contain a ribose ring, one with adenine attached to the first carbon atom and the other with nicotinamide at this position.
The nicotinamide moiety can be attached in two orientations to this anomeric carbon atom. Because of these two possible structures, the compound exists as two diastereomers, it is the β-nicotinamide diastereomer of NAD+, found in organisms. These nucleotides are joined together by a bridge of two phosphate groups through the 5' carbons. In metabolism, the compound donates electrons in redox reactions; such reactions involve the removal of two hydrogen atoms from the reactant, in the form of a hydride ion, a proton. The proton is released into solution, while the reductant RH2 is oxidized and NAD+ reduced to NADH by transfer of the hydride to the nicotinamide ring. RH2 + NAD+ → NADH + H+ + R; the midpoint potential of the NAD+/NADH redox pair is −0.32 volts, which makes NADH a strong reducing agent. The reaction is reversible, when NADH reduces another molecule and is re-oxidized to NAD+; this means the coenzyme can continuously cycle between the NAD+ and NADH forms without being consumed. In appearance, all forms of this coenzyme are white amorphous powders that are hygroscopic and water-soluble.
The solids are stable. Solutions of NAD+ are colorless and stable for about a week at 4 °C and neutral pH, but decompose in acids or alkalis. Upon decomposition, they form products. Both NAD+ and NADH absorb ultraviolet light because of the adenine. For example, peak absorption of NAD+ is at a wavelength of 259 nanometers, with an extinction coefficient of 16,900 M−1cm−1. NADH absorbs at higher wavelengths, with a second peak in UV absorption at 339 nm with an extinction coefficient of 6,220 M−1cm−1; this difference in the ultraviolet absorption spectra between the oxidized and reduced forms of the coenzymes at higher wavelengths makes it simple to measure the conversion of one to another in enzyme assays – by measuring the amount of UV absorption at 340 nm using a spectrophotometer. NAD+ and NADH differ in their fluorescence. NADH in solution has an emission peak at 460 nm and a fluorescence lifetime of 0.4 nanoseconds, while the oxidized form of the coenzyme does not fluoresce. The properties of the fluorescence signal changes when NADH binds to proteins, so these changes can be used to measure dissociation constants, which are useful in the study of enzyme kinetics.
These changes in fluorescence are used to measure changes in the redox state of living cells, through fluorescence microscopy. In rat liver, the total amount of NAD+ and NADH is 1 μmole per gram of wet weight, about 10 times the concentration of NADP+ and NADPH in the same cells; the actual concentration of NAD+ in cell cytosol is harder to measure, with recent estimates in animal cells ranging around 0.3 mM, 1.0 to 2.0 mM in yeast. However, more than 80% of NADH fluorescence in mitochondria is from bound form, so the concentration in solution is much lower. Data for other compartments in the cell are limited, although in the mitochondrion the concentration of NAD+ is similar to that in the cytosol; this NAD+ is carried into the mitochondrion by a specific membrane transport protein, since the coenzyme cannot diffuse across membranes. The balance between the oxidized and reduced forms of nicotinam
Deoxyribonucleic acid is a molecule composed of two chains that coil around each other to form a double helix carrying the genetic instructions used in the growth, development and reproduction of all known organisms and many viruses. DNA and ribonucleic acid are nucleic acids; the two DNA strands are known as polynucleotides as they are composed of simpler monomeric units called nucleotides. Each nucleotide is composed of one of four nitrogen-containing nucleobases, a sugar called deoxyribose, a phosphate group; the nucleotides are joined to one another in a chain by covalent bonds between the sugar of one nucleotide and the phosphate of the next, resulting in an alternating sugar-phosphate backbone. The nitrogenous bases of the two separate polynucleotide strands are bound together, according to base pairing rules, with hydrogen bonds to make double-stranded DNA; the complementary nitrogenous bases are divided into two groups and purines. In DNA, the pyrimidines are cytosine. Both strands of double-stranded DNA store the same biological information.
This information is replicated as and when the two strands separate. A large part of DNA is non-coding, meaning that these sections do not serve as patterns for protein sequences; the two strands of DNA are thus antiparallel. Attached to each sugar is one of four types of nucleobases, it is the sequence of these four nucleobases along the backbone. RNA strands are created using DNA strands as a template in a process called transcription. Under the genetic code, these RNA strands specify the sequence of amino acids within proteins in a process called translation. Within eukaryotic cells, DNA is organized into long structures called chromosomes. Before typical cell division, these chromosomes are duplicated in the process of DNA replication, providing a complete set of chromosomes for each daughter cell. Eukaryotic organisms store most of their DNA inside the cell nucleus as nuclear DNA, some in the mitochondria as mitochondrial DNA, or in chloroplasts as chloroplast DNA. In contrast, prokaryotes store their DNA only in circular chromosomes.
Within eukaryotic chromosomes, chromatin proteins, such as histones and organize DNA. These compacting structures guide the interactions between DNA and other proteins, helping control which parts of the DNA are transcribed. DNA was first isolated by Friedrich Miescher in 1869, its molecular structure was first identified by Francis Crick and James Watson at the Cavendish Laboratory within the University of Cambridge in 1953, whose model-building efforts were guided by X-ray diffraction data acquired by Raymond Gosling, a post-graduate student of Rosalind Franklin. DNA is used by researchers as a molecular tool to explore physical laws and theories, such as the ergodic theorem and the theory of elasticity; the unique material properties of DNA have made it an attractive molecule for material scientists and engineers interested in micro- and nano-fabrication. Among notable advances in this field are DNA origami and DNA-based hybrid materials. DNA is a long polymer made from repeating units called nucleotides.
The structure of DNA is dynamic along its length, being capable of coiling into tight loops and other shapes. In all species it is composed of two helical chains, bound to each other by hydrogen bonds. Both chains are coiled around the same axis, have the same pitch of 34 angstroms; the pair of chains has a radius of 10 angstroms. According to another study, when measured in a different solution, the DNA chain measured 22 to 26 angstroms wide, one nucleotide unit measured 3.3 Å long. Although each individual nucleotide is small, a DNA polymer can be large and contain hundreds of millions, such as in chromosome 1. Chromosome 1 is the largest human chromosome with 220 million base pairs, would be 85 mm long if straightened. DNA does not exist as a single strand, but instead as a pair of strands that are held together; these two long strands coil in the shape of a double helix. The nucleotide contains both a segment of the backbone of a nucleobase. A nucleobase linked to a sugar is called a nucleoside, a base linked to a sugar and to one or more phosphate groups is called a nucleotide.
A biopolymer comprising multiple linked nucleotides is called a polynucleotide. The backbone of the DNA strand is made from alternating sugar residues; the sugar in DNA is 2-deoxyribose, a pentose sugar. The sugars are joined together by phosphate groups that form phosphodiester bonds between the third and fifth carbon atoms of adjacent sugar rings; these are known as the 3′-end, 5′-end carbons, the prime symbol being used to distinguish these carbon atoms from those of the base to which the deoxyribose forms a glycosidic bond. When imagining DNA, each phosphoryl is considered to "belong" to the nucleotide whose 5′ carbon forms a bond therewith. Any DNA strand therefore has one end at which there is a phosphoryl attached to the 5′ carbon of a ribose and another end a
DNA ligase is a specific type of enzyme, a ligase, that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. It plays a role in repairing single-strand breaks in duplex DNA in living organisms, but some forms may repair double-strand breaks. Single-strand breaks are repaired by DNA ligase using the complementary strand of the double helix as a template, with DNA ligase creating the final phosphodiester bond to repair the DNA. DNA ligase is used in both DNA DNA replication. In addition, DNA ligase has extensive use in molecular biology laboratories for recombinant DNA experiments. Purified DNA ligase is used in gene cloning to join DNA molecules together to form recombinant DNA; the mechanism of DNA ligase is to form two covalent phosphodiester bonds between 3' hydroxyl ends of one nucleotide, with the 5' phosphate end of another. Two ATP molecules are consumed for each phosphodiester bond formed. AMP is required for the ligase reaction, which proceeds in four steps: Reorganization of activity site such as nicks in DNA segments or Okazaki fragments etc.
Adenylation of a lysine residue in the active center of the enzyme, pyrophosphate is released. Ligase will work with blunt ends, although higher enzyme concentrations and different reaction conditions are required; the E. coli DNA ligase is encoded by the lig gene. DNA ligase in E. coli, as well as most prokaryotes, uses energy gained by cleaving nicotinamide adenine dinucleotide to create the phosphodiester bond. It does not ligate blunt-ended DNA except under conditions of molecular crowding with polyethylene glycol, cannot join RNA to DNA efficiently; the activity of E. coli DNA ligase can be enhanced by DNA polymerase at the right concentrations. Enhancement only works when the concentrations of the DNA polymerase 1 are much lower than the DNA fragments to be ligated; when the concentrations of Pol I DNA polymerases are higher, it has an adverse effect on E. coli DNA ligase The DNA ligase from bacteriophage T4. The T4 ligase is the most-commonly used in laboratory research, it can ligate either cohesive or blunt ends of DNA, oligonucleotides, as well as RNA and RNA-DNA hybrids, but not single-stranded nucleic acids.
It can ligate blunt-ended DNA with much greater efficiency than E. coli DNA ligase. Unlike E. coli DNA ligase, T4 DNA ligase cannot utilize NAD and it has an absolute requirement for ATP as a cofactor. Some engineering has been done to improve the in vitro activity of T4 DNA ligase. A typical reaction for inserting a fragment into a plasmid vector would use about 0.01 to 1 units of ligase. The optimal incubation temperature for T4 DNA ligase is 16 °C. In mammals, there are four specific types of ligase. DNA ligase I: ligates the nascent DNA of the lagging strand after the Ribonuclease H has removed the RNA primer from the Okazaki fragments. DNA ligase III: complexes with DNA repair protein XRCC1 to aid in sealing DNA during the process of nucleotide excision repair and recombinant fragments. Of the all known mammalian DNA ligases, only Lig III has been found to be present in mitochondria. DNA ligase IV: complexes with XRCC4, it catalyzes the final step in the non-homologous end joining DNA double-strand break repair pathway.
It is required for VJ recombination, the process that generates diversity in immunoglobulin and T-cell receptor loci during immune system development. DNA ligase II: appears to be used in repair, it is formed by alternative splicing of a proteolytic fragment of DNA ligase III and does not have its own gene, therefore it is considered to be identical to DNA ligase III. DNA ligase from eukaryotes and some microbes uses adenosine triphosphate rather than NAD. Derived from a thermophilic bacterium, the enzyme is stable and active at much higher temperatures than conventional DNA ligases, its half-life is 48 hours at 65°C and greater than 1 hour at 95°C. Ampligase DNA Ligase has been shown to be active for at least 500 thermal cycles or 16 hours of cycling.10 This exceptional thermostability permits high hybridization stringency and ligation specificity. There are at least three different units used to measure the activity of DNA ligase: Weiss unit - the amount of ligase that catalyzes the exchange of 1 nmole of 32P from inorganic pyrophosphate to ATP in 20 minutes at 37°C.
This is the one most used. Modrich-Lehman unit - this is used, one unit is defined as the amount of enzyme required to convert 100 nmoles of dn to an exonuclease-III resistant form in 30 minutes under standard conditions. Many commercial suppliers of ligases use an arbitrary unit based on the ability of ligase to ligate cohesive ends; these units are more subjective than quantitative and lack precision. DNA ligases have become indispensable tools in modern molecular biology research for generating recombinant DNA sequences. For example, DNA ligases are used with restriction enzymes to insert DNA fragments genes, into plasmids. Controlling the optimal temperature is a vital aspect of performing effi