• The activity of E. coli DNA ligase can be enhanced by DNA polymerase at the right concentrations. (wikipedia.org)
  • Enhancement only works when the concentrations of the DNA polymerase 1 are much lower than the DNA fragments to be ligated. (wikipedia.org)
  • The replisome contains activities that separate the strands and hold them apart for synthesis by the replisome version of DNA polymerase, called DNA polymerase III in bacteria. (blogspot.com)
  • Deoxyribonucleic Acid (DNA) Recall that the replisome contains a DNA polymerase III holoenzyme dimer with two core complexes that can catalyze polymerization. (blogspot.com)
  • Each primer is extended from its 3′ end by DNA polymerase I to form an Okazaki fragment, as shown in the Figure. (blogspot.com)
  • The use of short RNA primers gets around the limitation imposed by the mechanism of DNA polymerase, namely, that it cannot initiate DNA synthesis de novo . (blogspot.com)
  • The primers are synthesized by a DNA dependent RNA polymerase enzyme called primase-the product of the dna G gene in E. coli . (blogspot.com)
  • The primosome, along with DNA polymerase III, is part of the replisome. (blogspot.com)
  • DNA polymerase III catalyzes synthesis of DNA in the 5′ → 3′ direction by extending each short RNA primer. (blogspot.com)
  • The steps are carried out by the combined action of DNA polymerase I and DNA ligase. (blogspot.com)
  • The LIC method uses T4 DNA polymerase to create specific 14 to 21 nucleotide single-stranded overhangs on the pLATE vectors and DNA inserts. (thermofisher.com)
  • T4 DNA polymerase has two enzymatic activities: 5'→3' polymerase activity and 3'→5' exonuclease activity. (thermofisher.com)
  • The exonuclease activity removes nucleotides from the 3' ends of the DNA while the polymerase activity restores the chain using dNTPs and the complementary DNA strand as a template. (thermofisher.com)
  • The final phosphodiester linkage between the 5'-phosphate group on the DNA chain synthesized by DNA polymerase III and the 3'-hydroxyl group on the chain made by DNA polymerase I is catalyzed by DNA ligase. (microbenotes.com)
  • DNA ends prepared by PCR for cloning may have a 3´ addition of a single adenine (A) residue as a result of amplification using a Taq DNA Polymerase (e.g. (bitesizebio.com)
  • Taq DNA Polymerase with Standard Taq Buffer, NEB #M0273 ) or have blunt ends if a high-fidelity DNA polymerase (e.g. (bitesizebio.com)
  • The fusion protein (PfuDBDlig-TaqS) was found to have enhanced processivity as a result of the conversion of the Taq DNA polymerase from a relatively low processive. (mostwiedzy.pl)
  • Lac DNA, RNA polymerase and cyclic AMP receptor protein, cyclic AMP, lac repressor and inducer are the essential elements for controlled lac transcription. (wikidata.org)
  • Purified DNA ligase is used in gene cloning to join DNA molecules together to form recombinant DNA. (wikipedia.org)
  • The E. coli DNA ligase is encoded by the lig gene. (wikipedia.org)
  • In both cases, the responsible gene was pifC , known also to be involved in FI of IncP plasmids and Agrobacterium T-DNA transfer to plant cells. (frontiersin.org)
  • It was also discovered that the R388 gene osa , which affects T-DNA transfer, also prevented conjugation of IncP-1/MOB P11 plasmids represented by plasmids RP4 and R751. (frontiersin.org)
  • Thermo Scientific aLICator™ LIC Cloning and Expression System is designed for fast and efficient ligation independent cloning and tight regulation of gene expression in E. coli . (thermofisher.com)
  • The tightly regulated expression and fast, efficient directional cloning makes the aLICator LIC Cloning and Expression System the best choice for routine and toxic gene cloning and expression in E. coli . (thermofisher.com)
  • DNA and Gene Organization of Phage Lambda 3. (biologydiscussion.com)
  • The codon table used was the FreqB table from " Design Parameters to Control Synthetic Gene Expression in Escherichia coli ", which was used because it was shown to express proteins well and was recommended to us by JCVI. (freegenes.org)
  • We establish a microbial biosynthetic pathway for PAA production from the simple substrate phenylalanine in E. coli with heterologous coexpression of aminotransferase (ARO8), keto acid decarboxylase (KDC) and aldehyde dehydrogenase H (AldH) gene. (springeropen.com)
  • In this study, we establish a microbial biosynthetic pathway for PAA production from the simple substrate phenylalanine through overexpression of an aminotransferase gene ARO8, a keto acid decarboxylase gene KDC from S. cerevisiae and an aldehyde dehydrogenase H gene aldH from E. coli in E. coli (Fig. 1 ). (springeropen.com)
  • This study evaluated the effect of matrix metalloproteinases-2 (MMP-2) gene silencing using MMP-2 shRNA expression plasmids (pMMP-2) on IR induced cytotoxicity and DNA damage by MTT, dead green, γH2AX and comet assays in human normal dermal fibroblasts (HDFs) and MCF-7 human breast cancer cells. (biomedcentral.com)
  • The aim of the present investigation is to study the effect of MMP-2 gene silencing by transfection of MMP-2 shRNA expression plasmids (pMMP-2on radiation-induced DNA damage in human normal dermal fibroblasts (HDFs) and MCF-7 cells. (biomedcentral.com)
  • The resulting novel recombinant gene was cloned and expressed in E. coli. (mostwiedzy.pl)
  • We also offer solutions for automation, site-directed mutagenesis, as well as your favorite restriction enzyme, ligase or competent cell products. (neb.com)
  • 5X enzyme concentration over standard T4 DNA Ligase, good for increasing reaction rates or for bulk users. (neb.com)
  • DNA ligase is a type of enzyme that facilitates the joining of DNA strands together by catalyzing the formation of a phosphodiester bond. (wikipedia.org)
  • Ligase will also work with blunt ends, although higher enzyme concentrations and different reaction conditions are required. (wikipedia.org)
  • Derived from a thermophilic bacterium, the enzyme is stable and active at much higher temperatures than conventional DNA ligases. (wikipedia.org)
  • DNA ligase is an enzyme that catalyzes the ligation reaction, ligation is the process of joining various types of DNA fragments. (microbenotes.com)
  • The enzyme DNA ligases were firstly isolated from E. coli in 1967, and several DNA ligases were discovered in 1967and 1968. (microbenotes.com)
  • DNA ligases are classified into two categories based on source, the enzyme requires either ATP or NAD + as a cofactor. (microbenotes.com)
  • The enzyme found in Haemophilus influenza consists of 268 amino acids, and larger cellular ligases such as human DNA ligase I consist of 912 amino acids, and IV consist of 844 amino acids. (microbenotes.com)
  • T4 DNA Ligase is a ligation enzyme that can be used to join DNA fragments by catalyzing the formation of phosphodiester bonds between juxtaposed 5' phosphate and 3' hydroxyl termini in double-stranded DNA using ATP as a coenzyme. (takarabio.com)
  • Properties of the enzyme-adenylate intermediate in the polynucleotide ligase reaction. (takarabio.com)
  • E. coli DNA Ligase is a NAD + -dependent enzyme that catalyzes the formation of phosphodiester bonds between complementary 3'-hydroxyl and 5'-phosphoryl termini of dsDNA. (mclab.com)
  • The enzyme joins blunt ends and cohesive ends as well as repair single stranded nicks in duplex DNA. (mclab.com)
  • The enzyme joins blunt end and cohesive end termini as well as repair single stranded nicks in duplex DNA. (mclab.com)
  • The 12 nucleotides of cohesive ends and process of circularization are shown in Fig. 18.11B-C. The events of circularization occurs after injection of phage DNA into E.coli cell where the bacterial enzyme, i.e. (biologydiscussion.com)
  • Which enzyme catalyzes the elongation of DNA molecules? (r4r.in)
  • 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. (wikipedia.org)
  • DNA ligase I: ligates the nascent DNA of the lagging strand after the Ribonuclease H has removed the RNA primer from the Okazaki fragments. (wikipedia.org)
  • DNA ligase III: complexes with DNA repair protein XRCC1 to aid in sealing DNA during the process of nucleotide excision repair and recombinant fragments. (wikipedia.org)
  • The short pieces of lagging-strand DNA are named Okazaki fragments in honor of their discoverer, Reiji Okazaki. (blogspot.com)
  • Okazaki fragments are eventually joined to produce a continuous strand of DNA. (blogspot.com)
  • The reaction proceeds in three steps: removal of the RNA primer, synthesis of replacement DNA, and sealing of the adjacent DNA fragments. (blogspot.com)
  • T4 DNA Ligase catalyzes the formation of phosphodiester bonds between double-stranded DNA fragments with 3'-OH and 5'-phosphate ends, in the presence of ATP. (takarabio.com)
  • Since the early 1970s, molecular cloning has used restriction enzymes to excise fragments from source DNA and to linearize plasmid vectors. (bitesizebio.com)
  • This step is important as it can drastically improve your cloning by separating your DNA from unwanted parent vectors and/or other DNA fragments. (bitesizebio.com)
  • NAD(+)-dependent DNA ligases (LigA) are ubiquitous in bacteria and essential for growth. (cornell.edu)
  • 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 (a bacteriophage that infects Escherichia coli bacteria). (wikipedia.org)
  • Bacteria display a series of mechanisms to control conjugative DNA transfer, as they do for any other physiological process ( Getino and de la Cruz, 2017 ). (frontiersin.org)
  • Whereas eukaryotic DNA ligases are ATP-dependent bacteria, archaea and virus DNA ligases are both NAD+ and ATP dependent. (microbenotes.com)
  • While some bacteria, archaea, and viruses, DNA ligases are both NAD+ and ATP dependent. (microbenotes.com)
  • The antimicrobial activity of purified stomoxyn-zy assessment indicated that, it had an in vitro fungcidal and bacteriocidal effect against fungi (Magnaporthe grisea), Gram-positive bacteria (staphylococcus aureus) and the Gram-negative bacteria (E. coli). (ukessays.com)
  • 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. (wikipedia.org)
  • As the replication fork progresses, the parental DNA is unwound, and more and more single-stranded DNA becomes exposed. (blogspot.com)
  • About once every second, primase catalyzes the synthesis of a short RNA primer using this single-stranded DNA as a template. (blogspot.com)
  • Both blunt and cohesive end DNA ligation, as well as single-stranded nick repair of DNA, RNA and DNA/RNA, are possible using the T4 DNA ligase. (takarabio.com)
  • Substrates include single-stranded RNA and DNA as well as dinucleoside pyrophosphates. (mclab.com)
  • Unlike T4 RNA Ligase 1, T4 RNA Ligase 2 is much more active joining nicks on double stranded RNA than on joining the ends of single stranded RNA. (mclab.com)
  • Background SSB (single-stranded DNA-binding) proteins play an essential role in all living cells and viruses, as they are involved in processes connected with ssDNA metabolism. (mostwiedzy.pl)
  • DNA ligase joins the DNA molecule covalently by catalyzing the formation of phosphodiester bonds between adjacent nucleotides. (microbenotes.com)
  • one successful approach, for example, tested T4 DNA ligase fused to several alternative DNA binding proteins and found that the constructs with either p50 or NF-kB as fusion partners were over 160% more active in blunt-end ligations for cloning purposes than wild type T4 DNA ligase. (wikipedia.org)
  • The pLATE bacterial expression vectors are designed for high levels of target protein expression in concert with minimal background (uninduced) expression, which permits expression of proteins that are toxic to E. coli cells. (thermofisher.com)
  • One of the most versatile defense mechanisms against the accumulation of DNA damage is nucleotide excision repair, in which, among others, the Xeroderma pigmentosum group C (XPC) and group A (XPA) proteins are involved. (aacrjournals.org)
  • A closed, ring-shaped conformation of ligase catalyzes a DNA end-joining reaction that is strongly stimulated by PCNA. (rcsb.org)
  • It catalyzes the final step in the non-homologous end joining DNA double-strand break repair pathway. (wikipedia.org)
  • T3 DNA Ligase catalyzes the formation of a phosphodiester bond between a 5′ phosphate and a 3′ hydroxyl terminus in duplex DNA. (mclab.com)
  • T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA using ATP as a cofactor. (mclab.com)
  • T4 RNA Ligase 1 catalyzes the ligation of a 5' phosphoryl-terminated nucleic acid donor to a 3' hydroxyl-terminated nucleic acid acceptor through the formation of a 3'→5' phosphodiester bond, with hydrolysis of ATP to AMP and PP i . (mclab.com)
  • Taq DNA Ligase catalyzes the formation of a phosphodiester bond in duplex DNA containing adjacent 5'-phosphoryl and 3'-hydroxyl termini, using NAD+ as a cofactor. (mclab.com)
  • It does not ligate blunt-ended DNA except under conditions of molecular crowding with polyethylene glycol, and cannot join RNA to DNA efficiently. (wikipedia.org)
  • It can also ligate blunt-ended DNA with much greater efficiency than E. coli DNA ligase. (wikipedia.org)
  • It can ligate either cohesive end or blunt ends of DNA, RNA -DNA hybrid, and also RNA. (microbenotes.com)
  • LigA envelopes the DNA within a protein clamp. (cornell.edu)
  • Therefore, molecular docking was also used in the present study to simulate the binding of cordycepin with different antibacterial targets, such as β-ketoacyl-(acyl-carrier-protein) synthase III (FabH), DNA gyrase B and thymidylate kinase, in order to identify the most likely antimicrobial mechanism of cordycepin. (spandidos-publications.com)
  • DNA sequence alignment analysis and the forecast-analysis of protein structure indicated that these seven antimicrobial peptide genes were not homologous to any other antimicrobial peptide genes in GenBank. (ukessays.com)
  • The head is made of protein of several types and contains a 46,500 bp long genomic (g) DNA. (biologydiscussion.com)
  • The phage λ contains double stranded circular DNA of about 17 µm in length packed in protein head of capsid. (biologydiscussion.com)
  • This open-to-closed switch in the conformation of DNA ligase is accommodated by a malleable interface with PCNA that serves as an efficient platform for DNA ligation. (rcsb.org)
  • Salt-T4 DNA Ligase is insensitive to salt carried over from other reaction components (vector or insert DNA) and allows ligation reactions to proceed in alternative reaction buffers with higher levels of salt (e.g. (neb.com)
  • 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 (94 °C/80 °C) or 16 hours of cycling.10 This exceptional thermostability permits extremely high hybridization stringency and ligation specificity. (wikipedia.org)
  • DNA ends prepared for cloning by restriction digest are ready for ligation, assuming the ends are compatible (that is, they have complementary overhangs or are blunt). (bitesizebio.com)
  • Successful digestion of your DNA should be confirmed on an agarose gel prior to ligation. (bitesizebio.com)
  • Salt-T4 DNA Ligase will join cohesive end termini at salt concentrations as high as 300 mM without any loss in activity. (neb.com)
  • DNA ligase II: A purification artifact resulting from proteolytic degradation of DNA ligase III. (wikipedia.org)
  • DNA ligase II: purification or excision repair of alternatively spliced form of DNA ligase III found in non-dividing cells. (microbenotes.com)
  • Direct PCR (dPCR) is a technique of DNA amplification straight from an animal or plant tissue pattern with out performing DNA isolation and purification steps. (gruenebundestag.de)
  • Adding template-switching primer, DNA ligase, or column purification of double-stranded cDNA does not improve the fidelity of amplification. (biomedcentral.com)
  • The size of DNA ligases is different from organism to organism. (microbenotes.com)
  • The resulting recombinant vector is then used to transform an E. coli host into a recombinant organism, expressing the exogenous DNA. (bitesizebio.com)
  • A typical reaction for inserting a fragment into a plasmid vector would use about 0.01 (sticky ends) to 1 (blunt ends) units of ligase. (wikipedia.org)
  • Domain la is required for the reaction of ligase with NAD + to form the ligase-AMP intermediate. (microbenotes.com)
  • Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB ® has the solution for you. (neb.com)
  • It ligates DNA during the process of nucleotide excision repair and recombinant fragment. (microbenotes.com)
  • The use of transposase (Tn5) to fragment ds DNA and integrate sequencing adapters in a single step - so called tagmentation - was first described by Adey et al. (biomedcentral.com)
  • ATP-dependent and NAD+ dependent ligases have a catalytic core. (microbenotes.com)
  • Many DNA ligases also contain a non-catalytic domain that is conserved, but the function of this domain is unknown. (microbenotes.com)
  • NAD + dependent ligases have a central ligase catalytic core which is composed of oligonucleotide binding fold domain and nucleotidyltransferase domain. (microbenotes.com)
  • Of the all known mammalian DNA ligases, only Lig III has been found to be present in mitochondria. (wikipedia.org)
  • DNA ligase III: It is the only mammalian ligase that is found in mitochondria. (microbenotes.com)
  • DNA ligase IV: homologous recombination repair and important in immune cell development and other activities critical to a normal cell. (microbenotes.com)
  • Construction of metabolically engineered E. coli strains with high stoichiometric yield of inositol is desirable. (biomedcentral.com)
  • The metabolically engineered E. coli strains represent a promising method for future inositol production. (biomedcentral.com)
  • CHIP is an E3 ligase, recognizing Hsp70-bound substrates via the TPR binding domain and catalyzing their ubiquitination via the U-box domain (hexagon). (elifesciences.org)
  • The i7 adapter was subsequently introduced by replacing the part of the i5 adapter that is not covalently linked to the tagmented DNA with an i7 adapter oligo. (biomedcentral.com)
  • Since the two strands of DNA are antiparallel, synthesis using one template strand occurs in the same direction as fork movement, but synthesis using the other template strand occurs in the direction opposite fork movement. (blogspot.com)
  • This limitation presents no difficulty for leading-strand synthesis since once DNA synthesis is under way nucleotides are continuously added to a growing chain. (blogspot.com)
  • When complexed with heterotrimeric PCNA, the DNA ligase binds to the PCNA3 subunit and ligase retains an open, extended conformation. (rcsb.org)
  • It plays a role in repairing single-strand breaks in duplex DNA in living organisms, but some forms (such as DNA ligase IV) may specifically repair double-strand breaks (i.e. a break in both complementary strands of DNA). (wikipedia.org)
  • 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 fully repair the DNA. (wikipedia.org)
  • Construction of four DNA sequences in biobrick format and pSB1A3 plasmid backbone which encode lock for medium RBS ( K175031 ), key for the lock of medium RBS ( K175032 ), lock for weak RBS ( K175029 ) or key for the lock of weak RBS ( K175030 ). (igem.org)
  • In mammals, there are four specific types of ligase. (wikipedia.org)
  • Unlike E. coli DNA ligase, T4 DNA ligase cannot utilize NAD and it has an absolute requirement for ATP as a cofactor. (wikipedia.org)
  • By breeding two of these mating-type engineered K. marxianus strains, we combined three complex traits-thermotolerance, lipid production, and facile transformation with exogenous DNA-into a single host. (asm.org)
  • It was found that PAA transformation yield was up to ~94% from phenylalanine in E. coli and there was no by-product phenethyl alcohol was detected. (springeropen.com)
  • DNA sliding clamps encircle DNA and provide binding sites for many DNA-processing enzymes. (rcsb.org)
  • DNA ligase belongs to the nucleotide transferase superfamily, including RNA ligases and mRNA capping enzymes. (microbenotes.com)
  • As with the standard procedure utilizing restriction enzymes, these recombinant constructions may then be used to transform an appropriate E. coli host. (bitesizebio.com)
  • Inositol was successfully biosynthesized after introducing two crucial enzymes: inositol-3-phosphate synthase (IPS) from Trypanosoma brucei , and inositol monophosphatase (IMP) from E. coli . (biomedcentral.com)
  • We designed our system with two different promotors, since we didn't know whether a high concentration of ''RIP'' will quorum quench the ''E. coli'' itself. (igem.org)
  • Transform into E. coli Top10 cells or equivalent. (freegenes.org)
  • To confirm that the improvement of CoQ10 production in E. coli Br-lyc was due to lycopene biosynthesis and not just geranylgeranyl diphosphate formation in the lycopene pathway, crtE was only introduced into E. coli Ba and E. coli Br strains. (ijbiotech.com)
  • Engineered route to stilbene biosynthesis in E. coli . (biomedcentral.com)
  • The biosynthesis of inositol from glucose in recombinant E. coli was optimized by metabolic engineering strategies. (biomedcentral.com)
  • A lycopene production pathway including geranylgeranyl diphosphate synthase (crtE), phytoene synthase (crtB), and phytoene desaturase (crtI) from Erwinia herbicola was constructed in two CoQ10-producing E. coli strains. (ijbiotech.com)
  • E. coli Ba and E. coli Br containing dds orthologs encoding for decaprenyl diphosphate synthase (Dds), respectively from Agrobacterium tumefaciens and Rhodobacter sphaeroides were transformed by the lycopene pathway resulting in E. coli Ba-lyc and E. coli Br-lyc. (ijbiotech.com)
  • DNA polymerases catalyze chain elongation exclusively in the 5′ → 3′ direction. (blogspot.com)
  • Zhou, W. , Richmond-Buccola, D. , Wang, Q. , and Kranzusch, P. J. (2022) Structural basis of human TREX1 DNA degradation and autoimmune disease . (cornell.edu)
  • Their distinctive substrate specificity and domain organization vis-a-vis human ATP-dependent ligases make them outstanding targets for anti-infective drug discovery. (cornell.edu)
  • The results of computational molecular docking analysis indicated that the interaction energy between cordycepin and NAD+‑dependent DNA ligase (LigA) was lower than that between cordycepin and other common antibacterial targets. (spandidos-publications.com)
  • Viral ATP-dependent DNA ligases are generally small in size. (microbenotes.com)
  • The lycopene pathway in E. coli Br-lyc interestingly resulted in a significant increase in CoQ10 production from 564 ± 28 to 989 ± 22 mg /g DCW. (ijbiotech.com)
  • Deletion mutants of simian virus 40 generated by enzymatic excision of DNA segments from the viral genome. (wikidata.org)
  • We have determined structures of Sulfolobus solfataricus DNA ligase and heterotrimeric PCNA separately by X-ray diffraction and in complex by small-angle X-ray scattering (SAXS). (rcsb.org)
  • After annealing, the LIC vector and insert are transformed into competent E. coli cells without the use of T4 DNA ligase. (thermofisher.com)
  • Take the steps to ensure your source DNA is free of such contaminants as nucleases and other unwanted enzymatic activities. (bitesizebio.com)
  • Energy is required for the joining of nicks (nicks lack the phosphodiester linkage between the nucleotide sequence) of DNA. (microbenotes.com)
  • The immunity operon products interact with DNA and decide whether the phage will initiate lytic cycle or lysogenic cycle. (biologydiscussion.com)
  • Accordingly, phaEC was constructed first in E. coli using a combination of a PCR method and the conventional "cut-and-paste" approach employing restriction endonucleases and DNA ligase. (usda.gov)