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DNA Ligases and Ligase Master Mixes STICK TOGETHER DNA Ligases and Ligase Master Mixes STICK TOGETHER be INSPIRED drive DISCOVERY stay GENUINE Stick together. DNA Ligases and Ligase Master Mixes from New England Biolabs With over 40 years of experience in the development and production of enzymes for molecular biology, NEB® now offers the most extensive selection of high-quality, and performance- optimized DNA ligases and ligase master mixes. Benefits of DNA Ligases and Ligase Master Mixes: • Fast ligations • Robust ligation efficiency • Industry standard for purity • Most extensive selection commercially available • New formulations optimized for your substrates • Convenient master mix formats • Highest fidelity Interested in tips to optimize your ligation experiments? Visit NEBStickTogether.com to view videos from NEB scientists. Choose the right DNA ligase product for your needs. NEB offers a variety of ligases for DNA research. All of these enzymes are recombinant, and all offer the quality and value you have come to expect from our products. While more than one ligase may work for your application, the following selection chart presents our recommendations for optimal performance. BLUNT/TA INSTANT NEBNEXT® LIGASE STICKY-END QUICK HIFI TAQ E. coli QUICK MASTER LIGASE ELECTRO- T4 DNA LIGATION™ T3 DNA T7 DNA DNA DNA TAQ DNA 9°N™ DNA LIGATION SPLINTR® MIX MASTER MIX LIGASE® LIGASE KIT LIGASE LIGASE LIGASE LIGASE LIGASE LIGASE MODULE LIGASE DNA APPLICATIONS Ligation of sticky ends «« ««« «« «« ««« «« «« « « « « Ligation of blunt ends ««« « «« «« ««« «« T/A cloning ««« « «« «« «« « « Electroporation ««« «« Ligation of sticky ends only ««« Repair of nicks in dsDNA «« «« «« ««« «« «« «« «« «« «« «« «« High complexity library cloning «« «« «« ««« «« Adaptor Ligation ««« «« «« « «« « ▲ Ligation Dependent DNA Sequence & SNP Detection (LCR, LDR, & related ««« «« «« methods) Ligation Dependent RNA Sequence & SNP Detection « ««« Ligation of adjacent ssDNA on an ssRNA Splint ««« NGS APPLICATIONS NGS Library Prep dsDNA-dsDNA ▲ ▲ ▲ ▲ (ligation) FEATURES Salt tolerance (> 2X that 4 of T4 DNA Ligase) Ligation in 15 min. or less 4 4 4 4 4 4 4 4 4 4 4 Master Mix Formulation 4 4 4 Thermostable 4 4 4 º Recombinant 4 4 4 4 4 4 4 4 4 4 4 4 4 ««« Optimal, recommended ligase for selected application «« Works well for selected application « Will perform selected application, but is not recommended ▲ Please consult the specific NGS protocol to determine the optimal enzyme for your needs Choose NEB DNA Ligases for quality and performance NEB ligases and ligase master mixes are manufactured Experience extreme purity Equivalent amounts of protein to the highest level of purity, and then rigorously with NEB's T4 DNA Ligase were loaded and silver stained using SilverXpress™. tested for optimal performance. NEB’s T4 DNA Ligase NEB NEB SUPPLIER M Lot 101 Lot 102 A B C D Marker M is NEB’s Broad Range has been referenced in peer-reviewed publications for Protein Marker (NEB #P7702). decades, and is renowned for its consistent quality. For details on quality controls performed, see www.neb.com/ligasequality. NEB's T4 DNA Ligase contains less contaminating activity T4 DNA Ligase from multiple Single-stranded suppliers was tested in reac- 120 Double-stranded blunt top tions containing a fluorescent Double-stranded blunt bottom labeled single stranded, double 3´ Overhang top stranded blunt, 3´overhang 100 3´ Overhang bottom or 5´overhang containing 5´ Overhang top oligonucleotides. The percent 5´ Overhang bottom degradation by contaminating 80 HC = High concentration nucleases is determined by cap- LC = Low concentration illary electrophoresis and peak analysis. The resolution is at the 60 single nucleotide level. adation (%) r Deg 40 20 0 1 2 1 2 1 2 1 2 1 1 2 3 1 1 2 1/HC 2/LC 2/HC 3/LC 3/HC 1 1 2 1/LC 2/HC A/HC B/HC C/HC A B C D E F G H I J K L Competitor Lots NEB Lots Ensure success with Blunt/TA Ligase Master Mix improves yields for ends Yields of final ligation product for that typically react slowly all reaction conditions using high formulations specific for concentration T4 DNA Ligase Nick 3' single-base overhang (NEB #M0202), The Quick Ligation Kit (NEB #M2200), and the Blunt/TA blunt-, T/A or sticky-ends Cohesive end 5' single-base overhang Ligase Master Mix (NEB #M0367). End architecture (blunt-, T/A or sticky-ends) Nick, cohesive end and cohesive end determines ligation efficiency. NEB offers ligases 3´ single-base overhang substrates were incubated for 15 minutes; the specifically optimized for either blunt or sticky 5´ single-base overhang was ends, including single-base overhangs. This incubated for 1 hour. increased efficiency gives you confidence that your cloning experiment will succeed the first time, every time. To learn more about how NEB's Blunt/TA Ligase Master Mix outperforms other ligases on difficult to ligate substrates, download our application notes at NEBStickTogether.com Properties of DNA Ligases This chart summarizes the activity information for NEB's DNA Ligases. For performance in various applications, see chart on other side. RECOMMENDED PRODUCT REACTION TEMP. LIGASE NUMBER (USEFUL RANGE) HEAT INACT. COFACTOR MAIN APPLICATIONS OTHER NOTES T4 DNA Ligase can ligate other substrates with reduced Ligation of nicks in dsDNA and joining efficiency, including: nicks containing mismatches and 1 or dsDNA fragments with complementary 2 nt gaps; dsDNA fragments with blunt ends and single-base 25°C overhangs > 2 bases in length. overhangs; and the 3´ end of RNA to a 5´pDNA when annealed T4 DNA Ligase M0202S/L (4–37°C) Y (65°C) ATP Routine cloning. to a DNA complement. Ligation of nicks in dsDNA and joining dsDNA fragments with complementary 5X enzyme concentration over standard T4 DNA Ligase, good for T4 DNA Ligase, 25°C overhangs > 2 bases in length. increasing reaction rates or for bulk users. See additional notes High Concentration M0202T/M (4–37°C) Y (65°C) ATP Routine cloning. for T4 DNA Ligase. Rapid ligation of dsDNA fragments Buffer contains PEG–do not heat inactivate or transform via 25°C with sticky or blunt ends. electroporation. Will also ligate nicks containing mismatches Quick Ligation™ Kit M2200 (4–37°C) N ATP Routine cloning. and 1 or 2 nt gaps with high efficiency. One-tube master mix format. Contains a proprietary Fastest ligation of blunt and single base enhancer that increases ligation yields. Contains PEG–do not overhang (T/A) substrates. T/A cloning, heat inactivate or transform via electroporation. Will also ligate 25°C Best choice for adaptor ligation in NGS nicks containing mismatches and 1 or 2 nt gaps with high Blunt/TA Ligase Master Mix M0367 (4–37°C) N ATP sequencing library preparation. efficiency. One tube master mix format. Contains a proprietary enhancer that increases ligation yields. Contains PEG–do not heat inactivate or transform via electroporation. Will also ligate Instant Sticky End Ligase 25°C Fastest ligation of dsDNA with sticky ends. nicks containing mismatches and 1 or 2 nt gaps with high Master Mix M0370 (4–37°C) N ATP Routine cloning. efficiency. Efficient ligation of nicks, sticky ends, and blunt or T/A ends when a PEG–free 25°C mixture is required. Suitable for direct Contains a proprietary enhancer that increases ligation yields. ElectroLigase® M0369 (4–37°C) Y (65°C) ATP transformation via electroporation. See also notes for T4 DNA Ligase. Buffer contains PEG–do not heat inactivate or transform via A salt-tolerant enzyme for the ligation of electroporation. Can be used with the PEG-free T4 DNA Ligase 25°C nicks, cohesive ends, and blunt ends in Buffer (NEB #B0202) with reduced activity. Can also ligate the 3´ T3 DNA Ligase M0317 (4–37°C) N ATP dsDNA. Routine cloning. end of RNA to 5´-pRNA when annealed to a DNA complement. Buffer contains PEG–do not heat inactivate or transform via Allows selective ligation of nicks and electroporation. High specificity for correctly base-paired nicks 25°C cohesive ends in dsDNA, not ligating if used with T4 DNA Ligase Buffer (NEB #B0202) instead of T7 DNA Ligase M0318 (4–37°C) N ATP blunt ends or single base overhangs. supplied T7 DNA Ligase Buffer. Selective ligation of nicks in dsDNA without significant joining of dsDNA 25°C fragments regardless of end type. E. coli DNA Ligase M0205 (4–37°C) Y (65°C) NAD cDNA synthesis. Used in some cDNA library preparation protocols. Thermostable NAD dependent ligase that ligates only nicks in dsDNA with the highest discrimination against mismatched bases. 60°C Useful for SNP detection through LDR and Use Thermostable Ligase Reaction Calculator to estimate HiFi Taq DNA Ligase M0647S (37–75°C) N NAD other ligation based detection methods. incubation temperature. Thermostable ATP-dependent ligase that ligates only nicks in dsDNA with a high discrimination against mismatched bases. 60°C Useful for LDR and other ligation-based 9°N™ DNA Ligase M0238 (45–70°C) N ATP detection methods. Thermostable NAD-dependent ligase that ligates only nicks in dsDNA with a high discrimination against mismatched bases. 60°C Useful for LDR and other ligation-based Taq DNA Ligase M0208 (37–75°C) N NAD detection methods. Used in Gibson Assembly® methods. Ligation of the 3´ end of DNA to a 5´-pDNA annealed to an RNA complement. Specific Ligates fully DNA substrates with high efficiency. Can also ligate 25°C detection of RNA sequences through the the 3´ end of RNA to 5´-pDNA when annealed to either DNA or SplintR® Ligase M0375 (4–37°C) Y (65°C) ATP ligation of complementary DNA probes. RNA complements. USA Choose from the widest selection New England Biolabs, Inc. Telephone (978) 927-5054 Toll Free (USA Orders) 1-800-632-5227 of ligases commercially available Toll Free (USA Tech) 1-800-632-7799 Fax (978) 921-1350 NEB offers 13 different DNA ligase products, more than any other supplier.
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