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2019-20 NEB Catalog Technical Reference

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2019-20 NEB Catalog Technical Reference Reference Appendix Technical Support – Featured Tools & for scientists, by scientists Resources As a partner to the scientific community, New England Biolabs is committed to Optimizing Restriction Enzyme Reactions providing top quality tools and scientific expertise. This philosophy still stands, 290 and has led to long-standing relationships with many of our fellow scientists. Performance Chart for NEB's commitment to scientists is the same regardless of whether or not they 293 Restriction Enzymes purchase product from NEB: their ongoing research is supported by our catalog, Troubleshooting Guide website and technical staff. 349 for Cloning NEB's technical support model is unique as it utilizes most of the scientists at NEB. Several of our product lines have designated technical support scientists 334 Methylation Sensitivity assigned to servicing customers in those application areas. Any questions regarding a product could be dealt with by one of the technical support Guidelines for scientists, the product manager who manufactures it, the product development 337 PCR Optimization scientist who optimizes it, or a researcher who uses the product in their daily Cleavage Close to the research. As such, customers are supported by scientists and often experts in 343 End of DNA Fragments the product or its application. To access technical support: 288 Online Interactive Tools Call 1-800-632-7799 (Monday – Friday: 9:00 am - 6:00 pm EST) Submit an online form at www.neb.com/techsupport View NEB TV Episode #22 Email [email protected] to learn more about our Technical Support program. International customers can contact a local NEB subsidiary or distributor. For more information see inside back cover. Visit the Tools & Resources tab at www.neb.com to find additional online tools, video tech tips and tutorials to help Learn more you in your research. about NEB's tech support program. 286 Online Interactive Tools 288 PCR, qPCR & RT-qPCR Databases, Mobile Apps & General Guidelines for PCR Optimization 337 Freezer Program Locator 289 PCR Troubleshooting Guide 338 Restriction Endonucleases Optimization Tips for Luna qPCR 339 Optimization Tips for Luna One-Step RT-qPCR 340 Optimizing Restriction Enzyme Reactions 290 Luna qPCR Troubleshooting Guide 341 Double Digestion 291 Luna One-Step RT-qPCR Troubleshooting Guide 342 Types of Restriction Enzymes 291 Cleavage Close to the End of DNA Fragments 343 Troubleshooting Guide 292 Activity of Restriction Enzymes in PCR Buffers 344–345 Performance Cloning Performance Chart for Restriction Enzymes 293–298 Getting Started with Molecular Cloning 346 Activity of Enzymes at 37°C 299 Traditional Cloning Quick Guide 347–348 Activity of DNA Modifying Enzymes in CutSmart Buffer 299 Troubleshooting Guide for Cloning 349–351 Tips for Avoiding Star Activity 300 Optimization Tips for Your Cloning Reaction 352–353 High-Fidelity (HF) Restriction Enzymes 301 Nucleic Acid Purification Reduced Star Activities of HF Enzymes 301 Troubleshooting Guide for DNA Cleanup Time-Saver Qualified Restriction Enzymes 302–303 & Plasmid Purification Using Monarch Kits 354 Isoschizomers/Recognition Sequences Choosing Sample Input Amounts When Using the Monarch Genomic DNA Purification Kit 355 Cross Index of Recognition Sequences 304–306 Troubleshooting Guide for Genomic DNA Palindromic Penta-nucleotide Purification Using Monarch Kits 356 Recognition Sequences 305 Choosing Sample Input Amounts When Using the Specificities Greater Than 6 Bases 305 Monarch Total RNA Miniprep Kit 357 Interrupted Palindromes 305–306 Troubleshooting Guide for Total RNA Extraction Multiple & Nonpalindromic & Purification using Monarch Kits 358 Recognition Sequences 306–309 Alphabetized List of NEB Recognition Sequences 309–310 Transformation Isoschizomers 311–327 Genetic Markers 359–360 Properties of Restriction Enzymes Enhancing Transformation Efficiency 361 Protein Expression with T7 Express Strains 362 Survival in a Reaction 328–329 Cleavage of Supercoiled DNA 329 NGS Library Preparation Generating New Cleavage Sites DNA/RNA Input Guidelines for NGS Library Prep 363 Recleavable Filled-in 5´ Overhangs 330 Protein Expression Recleavable Blunt Ends 331 Labeling with SNAP-tag Technology Compatible Cohesive Ends 332–333 Troubleshooting Guide 364 Frequencies of Restriction Sites in Methylation Sensitivity Sequenced DNAs 365–368 Dam, Dcm and CpG Methylation 334–336 DNA Maps 369–381 Odds and Ends General Molecular Biology Information 382–385 Index 386 One or more of these products are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc. For more information, please email us at [email protected]. The use of these products may require you to obtain additional third party intellectual property rights for certain applications. Your purchase, acceptance, and/or payment of and for NEB’s products is pursuant to NEB’s Terms of Sale at https://www.neb.com/support/terms-of-sale. NEB does not agree to and is not bound by any other terms or conditions, unless those terms and conditions have been expressly agreed to in writing by a duly authorized officer of NEB. 287 Online Interactive Tools Use the Tools & Resources tab at www.neb.com to access our growing selection of interactive technical tools. These tools can also be accessed directly in the footer of every web page. NEB scientists are often involved in the development of online tools that will aid in their research. We are now making these tools and in some circumstances, the source code, available for you to evaluate. To learn more, visit www.neb.com/NEBetaTools. Online Tools Competitor Cross-Reference Tool NEBcutter® V2.1 Use this tool to select another company's product and find out Identify the restriction sites within your DNA sequence using which NEB product is compatible. Choose either the product NEBcutter. Choose between Type II and commercially available name or catalog number from the available selections, and this Type III restriction enzymes to digest your DNA. NEBcutter will tool will identify the recommended NEB product. indicates cut frequency and methylation-state sensitivity. ® DNA Sequences and Maps Tool NEBioCalculator Use this tool for your scientific calculations and conversions With the DNA Sequences and Maps Tool, find the nucleotide for DNA and RNA. Options include conversion of mass to sequence files for commonly used molecular biology tools, moles, ligation amounts, conversion of OD to concentration, including plasmid, viral and bacteriophage vectors. dilution and molarity. Additional features include sgRNA template oligo design and qPCR library quantification. ® Double Digest Finder NEBcloner Use this tool to find the right products and protocols for each Use this tool to guide your reaction buffer selection step (digestion, end modification, ligation, transformation when setting up double digests, a common timesaving and mutagenesis) of your next traditional cloning experiment. procedure. Choosing the right buffers will help you to Also, find other relevant tools and resources to enable protocol avoid star activity and loss of product. optimization. ® Enzyme Finder NEBuilder Assembly Tool Use this tool to design primers for your DNA assembly Use this tool to select restriction enzymes by name, reaction, based on the entered fragment sequences and the sequence, overhang or type. Enter your sequence polymerase being used for amplification. using single letter code nomenclature, and Enzyme Finder will identify the right enzyme for the job. Glycan Analyzer PCR Fidelity Estimator Use this tool to interpret ultra or high pressure liquid Estimate the percentage of correct DNA copies (those chromatography (UPLC/HPLC) N-glycan profiles without base substitution errors) per cycle of PCR for following exoglycosidase digestions. selected DNA polymerases. NEB Golden Gate Assembly Tool PCR Selector Use this tool to assist with in silico DNA construct design for Use this tool to help select the right DNA polymerase Golden Gate DNA assembly. It enables the accurate design of for your PCR setup. Whether your amplicon is long, primers with appropriate Type IIS restriction sites and overlaps, complex, GC-rich or present in a single copy, the PCR quick import of sequences in many formats and export of the selection tool will identify the perfect DNA polymerase final assembly, primers and settings. for your reaction. NEBaseChanger® Tm Calculator NEBaseChanger can be used to design primers specific to Determine the optimal annealing temperature for your amplicon the mutagenesis experiment you are performing using the with our Tm Calculator. Simply input your DNA polymerase, Q5® Site-Directed Mutagenesis Kit. This tool will also calculate primer concentration and your primer sequence, and the Tm APPENDIX a recommended custom annealing temperature based on the Calculator will guide you to successful reaction conditions. sequence of the primers by taking into account any mismatches. NEBNext® Selector Thermostable Ligase Reaction Temperature Calculator Use this tool to guide you through the selection of NEBNext This tool will help you estimate an optimal reaction temperature reagents for next generation sequencing sample preparation. to minimize mismatch for thermostable ligation of two adjacent ssDNA probes annealed to a template. 288 www.neb.com · [email protected] TECHNICAL SUPPORT Online Resources Online Tools (continued) Read Coverage Calculator This tool allows for easy calculation of values associated with read coverage in NGS protocols. Additional Databases
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