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Query: EC:6.5.1.2 (DNA ligase)
 2,749 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

DNA ligases of bacteriophage T4 and T7 have been widely used in molecular biology for decades, but little is known about bacteriophage T3 DNA ligase. Here is the first report on the cloning, expression and biochemical characterization of bacteriophage T3 DNA ligase. The polyhistidine-tagged recombinant T3 DNA ligase was shown to be an ATP-dependent enzyme. The enzymatic activity was not affected by high concentration of monovalent cations up to 1 M, whereas 2 mM ATP could inhibit its activity by 50%. Under optimal conditions (pH 8.0, 0.5 mM ATP, 5 mM DTT, 1 mM Mg(2+) and 300 mM Na(+)), 1 fmol of T3 DNA ligase could achieve 90% ligation of 450 fmol of cohesive dsDNA fragments in 30 min. T3 DNA ligase was shown to be over 5-fold more efficient than T4 DNA ligase for ligation of cohesive DNA fragments, but less active for blunt-ended DNA fragments. Phylogenetic analysis showed that T3 DNA ligase is more closely related to T7 DNA ligase than to T4 DNA ligase.
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PMID:Characterization of bacteriophage T3 DNA ligase. 1511 38

This appendix describes the preparation of selected bacterial media and of buffers and reagents used in the manipulation of nucleic acids and proteins. Recipes for cell culture media and reagents are located elsewhere in the manual. RECIPES: Acids, concentrated stock solutions; Ammonium acetate, 10 M; Ammonium hydroxide, concentrated stock solution; ATP, 100 mM; BCIP, 5% (w/v); BSA (bovine serum albumin), 10% (100 mg/ml); Denhardt solution, 100x; dNTPs: dATP, dTTP, dCTP, and dGTP; DTT, 1 M; EDTA, 0.5 M (pH 8.0); Ethidium bromide solution; Formamide loading buffer, 2x; Gel loading buffer, 6x; HBSS (Hanks balanced salt solution); HCl, 1 M; HEPES-buffered saline, 2x; KCl, 1 M; LB medium; LB plates; Loading buffer; 2-ME, (2-mercaptoethanol)50 mM; MgCl(2), 1 M; MgSO(4), 1 M; NaCl, 5 M; NaOH, 10 M; NBT (nitroblue tetrazolium chloride), 5% (w/v); PCR amplification buffer, 10x; Phosphate-buffered saline (PBS), pH approximately 7.3; Potassium acetate buffer, 0.1 M; Potassium phosphate buffer, 0.1 M; RNase a stock solution (DNase-free), 2 mg/ml; SDS, 20%; SOC medium; Sodium acetate, 3 M; Sodium acetate buffer, 0.1 M; Sodium phosphate buffer, 0.1 M; SSC (sodium chloride/sodium citrate), 20x; SSPE (sodium chloride/sodium phosphate/EDTA), 20x; T4 DNA ligase buffer, 10x; TAE buffer, 50x; TBE buffer, 10x; TBS (Tris-buffered saline); TCA (trichloroacetic acid), 100% (w/v); TE buffer; Terrific broth (TB); TrisCl, 1 M; TY medium, 2x; Urea loading buffer, 2x.
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PMID:Common buffers, media, and stock solutions. 1842 17