{"product_id":"gb-clone-fast-t4-dna-ligase","title":"GB-Clone™ Fast T4 DNA Ligase","description":"\u003cp\u003e\u003cspan\u003eGB-Clone™ \u003c\/span\u003eFAST T4 DNA Ligase is designed for the efficient ligation of cohesive-ended DNA inserts\u003c\/p\u003e\n\u003cp\u003einto plasmid vectors in just 5-10 minutes (blunt-ended inserts in as little as 15-30 minutes).\u003cbr\u003eFAST T4 DNA Ligase catalyzes the joining of two strands of DNA between the\u003cbr\u003e5´-phosphate and the 3´-hydroxyl groups of adjacent nucleotides in either a\u003cbr\u003ecohesive-ended or blunt-ended configuration. The enzyme has also been shown to\u003cbr\u003ecatalyze the joining of RNA to either a DNA or RNA strand in a duplex molecule but will not\u003cbr\u003ejoin single-stranded nucleic acids.\u003c\/p\u003e\n\u003cp\u003eBiological Source: E. coli strain expressing a recombinant clone.\u003cbr\u003eMolecular Weight: 68kDa.\u003cbr\u003eRequirements: Mg2+, ATP and DTT. The optimum concentration of Mg2+ is 10mM. Mn2+ may be\u003cbr\u003esubstituted for Mg2+ but is only 25% as effective as Mg2+.\u003cbr\u003eInhibition: 50% inhibition by greater than 150mM NaCl (activity measured at nicks. Other inhibitors include 0.2M K+, Cs+, Li+, NH4+ and 1mM spermine.\u003cbr\u003eInactivation: Heat to 70°C for 10 minutes.\u003c\/p\u003e\n\u003cp\u003eContents: FAST T4 DNA Ligase (400 U\/μl)\u003cbr\u003e2 X Ligation Buffer\u003c\/p\u003e\n\u003cp\u003eWe recommend that 0.5-1 μl of the ligase is used per 10 μl ligation reaction. \u003c\/p\u003e\n\u003cp\u003eApplications\u003cbr\u003e Cloning of restriction fragments.\u003cbr\u003e Joining linkers and adapters to blunt-ended DNA\u003c\/p\u003e\n\u003cp\u003eUnit Definition\u003cbr\u003eOne unit of FAST T4 DNA Ligase is defined as the amount of enzyme required to give\u003cbr\u003e50% ligation of Hind III fragments of λ DNA (5´ DNA termini concentration of 0.12 μM, 300-\u003cbr\u003eμg\/ml) in a total reaction volume of 20 μl in 30 minutes at 16°C in 1x T4 DNA Ligase\u003cbr\u003eReaction Buffer.\u003cbr\u003e\u003cbr\u003eStorage： Store at -20℃\u003cbr\u003eStorage Buffer: 50 mM KCl, 10 mM Tris-HCl (pH 7.4), 0.1 mM EDTA, 1 mM DTT, 200 μg\/ml BSA and 50% glycerol.\u003c\/p\u003e\n\u003cp\u003e2X T4 DNA Ligase Buffer: 60mM Tris-HCl (pH 7.8), 20mM MgCl2, 20mM DTT, 2mM ATP and 10% PEG.\u003c\/p\u003e\n\u003cp\u003ePhysical Purity: The purity is ≥90% as judged by SDS-polyacrylamide gel electrophoresis with\u003cbr\u003eCoomassie® blue staining.\u003c\/p\u003e\n\u003cp\u003eStandard Applications\u003cbr\u003eWe recommend using a 1:1, 1:3 or 3:1 molar ratio of vector:insert DNA when cloning a\u003cbr\u003efragment into a plasmid vector. These ratios will vary with other types of vectors, for\u003cbr\u003eexample, cDNA and genomic cloning vectors. The following example illustrates the\u003cbr\u003econversion of molar ratios to mass ratios for a 3.0kb plasmid and a 0.5kb insert DNA\u003cbr\u003efragment.\u003c\/p\u003e\n\u003cp\u003eProtocol:\u003cbr\u003eThe following ligation reaction of a 3kb vector and a 0.5kb insert DNA uses a 1:1\u003cbr\u003evector:insert ratio. Typical ligation reactions use 100–200ng of vector DNA.\u003cbr\u003e1. Assemble the following reaction in a sterile microcentrifuge tube:\u003cbr\u003evector DNA 100ng\u003cbr\u003einsert DNA 17ng\u003cbr\u003e2 X Ligation Buffer 5μl\u003cbr\u003eT4 DNA Ligase 0.5–1μl\u003cbr\u003eNuclease-Free Water to final volume of 10μl\u003cbr\u003e2. Incubate the reaction at room temperature for 5-10 minutes for cohesive-ended\u003cbr\u003eligations, or 15-30 minutes for blunt-ended ligations.\u003cbr\u003eNotes:\u003cbr\u003e1. Ligation reactions performed using the 2 X Ligation Buffer do not need to be cleaned up\u003cbr\u003ebefore transformation.\u003cbr\u003e2. Concatamers may form as ligation products. The extent of concatamer formation\u003cbr\u003edepends on the vector:insert ratio, incubation temperature and incubation time. This\u003cbr\u003eshould be taken into account when screening transformants.\u003c\/p\u003e","brand":"Gene Bio Systems","offers":[{"title":"50 uL","offer_id":39766557130852,"sku":"GPL-01","price":102.95,"currency_code":"EUR","in_stock":true},{"title":"200 ul","offer_id":39766557163620,"sku":"GPL-02","price":284.95,"currency_code":"EUR","in_stock":true}],"thumbnail_url":"\/\/cdn.shopify.com\/s\/files\/1\/0558\/8588\/9636\/products\/329.jpg?v=1643677710","url":"https:\/\/www.genebiosystems.com\/en-de\/products\/gb-clone-fast-t4-dna-ligase","provider":"GeneBio ","version":"1.0","type":"link"}