DNA Cloning and Mapping and Mutagenesis
DNA Cloning and Mapping and Mutagenesis
New England Biolabs, Inc. NEB Golden Gate Assembly Kit (BsmBI-v2) – 100 reactions
The NEB Golden Gate Assembly Kit (BsmBI-v2) contains an optimized mix of BsmBI-v2 and T4 DNA Ligase. BsmBI-v2 has been engineered by NEB and outperforms BsmBI in Golden Gate Assemblies. Together these enzymes can direct the assembly of multiple inserts/modules and also single insert/library generation cloning with single insert(s) using the Golden Gate approach. The pGGAselect destination plasmid provides a backbone for your assembly. It has flanking recognition sites in the correct orientation for BsmBI-directed assemblies, and also BsaI- and BbsI-directed assemblies, enabling the destination plasmid to conveniently be used with the most commonly used Type IIS restriction enzymes used for Golden Gate Assembly. Please note that while general descriptions regarding Golden Gate Assembly use the BsmBI nomenclature, this kit and protocols feature the specific engineered form optimized for Golden Gate Assembly, BsmBI-v2.
New England Biolabs, Inc. NEB Golden Gate Enzyme Kit (BsaI-HFv2) – 100 reactions
The NEB Golden Gate Assembly Kit (BsaI-HFv2) contains an optimized mix of BsaI-HFv2 and T4 DNA Ligase. Together these enzymes can direct the assembly of multiple inserts/modules using the Golden Gate approach. Also included is the pGGAselectdestination plasmid, which provides a backbone for your assembly, features convenient restriction enzyme sites for subcloning, and has T7/SP6 promoter sequences to enable in vitro transcription. Please note that while general descriptions regarding Golden Gate Assembly use the BsaI nomenclature, this kit and protocols feature the specific engineered form optimized for Golden Gate Assembly, BsaI-HFv2.
New England Biolabs, Inc. NEBuilder HiFi DNA Assembly Master Mix – 50 reactions
NEBuilder HiFi DNA Assembly Master Mix was developed to improve the efficiency and accuracy of DNA assembly. It allows for seamless assembly of multiple DNA fragments, regardless of fragment length or end compatibility. Used to assemble either ss oligonucleotides or different sizes of DNA frags with varied overlaps. Has utility in synbio community, as well as those interested in one-step cloning of multiple fragments due to its ease of use, flexibility and simple MM format. The rxn includes different enzymes that work together in the same buffer: The exonuclease creates single-stranded 3' overhangs that facilitate the annealing of fragments that share complementarity at one end (the overlap region): a polymerase fills in gaps within each annealed fragment; DNA ligase seals nicks in the assembled DNA. End result is a ds fully sealed DNA molecule that can serve as template for PCR, RCA or a variety of other molecular biology applications, including direct transformation of E coli.
New England Biolabs, Inc. NEBNext End Repair Module – 100 reactions
This module is part of the original standard library prep workflow, which is longer than the NEBNext Ultra II DNA Library Prep Kit workflow, has more components, and requires more clean-up steps. It also requires higher input amounts: 1 - 5 µg of input DNA vs. 500 pg - 1 µg required for the Ultra II DNA workflow.
New England Biolabs, Inc. Lambda DNA – 1250 µg
Lambda DNA is a commonly used DNA substrate. It is duplex DNA is isolated from bacteriophage lambda (cI857ind 1 Sam 7). Lambda DNA is 48,502 base pairs in length.
New England Biolabs, Inc. Quick Ligation™ Kit – 30 reactions
The Quick Ligation Kit enables ligation of cohesive end or blunt end DNA fragments in 5 minutes at room temperature. (25C)
New England Biolabs, Inc. NEBuilder HiFi DNA Assembly Master Mix – 10 reactions
NEBuilder HiFi DNA Assembly Master Mix was developed to improve the efficiency and accuracy of DNA assembly. It allows for seamless assembly of multiple DNA fragments, regardless of fragment length or end compatibility. Used to assemble either ss oligonucleotides or different sizes of DNA frags with varied overlaps. Has utility in synbio community, as well as those interested in one-step cloning of multiple fragments due to its ease of use, flexibility and simple MM format. The rxn includes different enzymes that work together in the same buffer: The exonuclease creates single-stranded 3' overhangs that facilitate the annealing of fragments that share complementarity at one end (the overlap region): a polymerase fills in gaps within each annealed fragment; DNA ligase seals nicks in the assembled DNA. End result is a ds fully sealed DNA molecule that can serve as template for PCR, RCA or a variety of other molecular biology applications, including direct transformation of E coli.
New England Biolabs, Inc. NEBuilder HiFi DNA Assembly Cloning Kit – 10 reactions
NEBuilder HiFi DNA Assembly Master Mix was developed to improve the efficiency and accuracy of DNA assembly. This method allows for seamless assembly of multiple DNA fragments, regardless of fragment length or end compatibility. This method has been used to assemble either single-stranded oligonucleotides or different sizes of DNA fragments with varied overlaps (15-30 bp). It has utility for the synthetic biology community, as well as those interested in one-step cloning of multiple fragments due to its ease of use, flexibility and simple master-mix format. The cloning kit contains NEB 5-alpha Competent Cells.
New England Biolabs, Inc. EnGen Mutation Detection Kit – 25 reactions
The EnGen Mutation Detection Kit provides reagents for detection of on-target genome editing events. In the first step, targeted regions from cells whose genomes were targeted (i.e. CRISPR/Cas9, TALENs, Zinc-finger Nucleases) are amplified using Q5 Hot Start High-Fidelity 2X Master Mix. Upon denaturation and re-annealing, heteroduplexes are formed when mutations from insertions and deletions (indels) are present in the amplicon pool. In the second step, annealed PCR products are digested with EnGen T7 Endonuclease I, a structure-specific enzyme that will recognize mismatches larger than 1 base. Both strands of the DNA are cut when a mismatch is present, which results in the formation of smaller fragments. Analysis of the resulting fragments provides an estimate of the efficiency of the genome editing experiments. The EnGen Mutation Detection Kit includes a Control Template and Primer Mix that can be used as a control for the PCR reaction and T7 Endonuclease I digestion.
New England Biolabs, Inc. Q5 Site-Directed Mutagenesis Kit – 10 reactions
The Q5 Site-Directed Mutagenesis Kit enables rapid, site-specific mutagenesis of double-stranded plasmid DNA in less than 2 hours. The kit utilizes the robust Q5 Hot Start High-Fidelity DNA Polymerase along with custom mutagenic primers to create insertions, deletions and substitutions in a wide variety of plasmids. After PCR, the amplified material is added directly to a unique Kinase-Ligase-DpnI (KLD) enzyme mix for rapid (5 minutes), room temperature circularization and template removal. Transformation into high-efficiency chemically-competent E. coli, ensures robust results with plasmids up to at least 20 kb in length.
New England Biolabs, Inc. NEBuilder HiFi DNA Assembly Bundle for Large Fragments – 20 reactions
NEB recommends NEB 10-beta Competent E. coli , High Efficiency for assemblies larger than 15 kb. NEBuilder HiFi DNA Assembly Bundle for Large Fragments offers a discount for purchasing these competent cells with the NEBuilder HiFi DNA Assembly Master Mix. NEBuilder HiFi DNA Assembly Master Mix was developed to improve the efficiency and accuracy of DNA assembly. This method allows for seamless assembly of multiple DNA fragments, regardless of fragment length or end compatibility. This method has been used to assemble either single-stranded oligonucleotides or different sizes of DNA fragments with varied overlaps (15-30 bp). It has utility for the synthetic biology community, as well as those interested in one-step cloning of multiple fragments due to its ease of use, flexibility and simple master-mix format.
New England Biolabs, Inc. NEB Golden Gate Assembly Kit (BsmBI-v2) – 20 reactions
The NEB Golden Gate Assembly Kit (BsmBI-v2) contains an optimized mix of BsmBI-v2 and T4 DNA Ligase. BsmBI-v2 has been engineered by NEB and outperforms BsmBI in Golden Gate Assemblies. Together these enzymes can direct the assembly of multiple inserts/modules and also single insert/library generation cloning with single insert(s) using the Golden Gate approach. The pGGAselect destination plasmid provides a backbone for your assembly. It has flanking recognition sites in the correct orientation for BsmBI-directed assemblies, and also BsaI- and BbsI-directed assemblies, enabling the destination plasmid to conveniently be used with the most commonly used Type IIS restriction enzymes used for Golden Gate Assembly. Please note that while general descriptions regarding Golden Gate Assembly use the BsmBI nomenclature, this kit and protocols feature the specific engineered form optimized for Golden Gate Assembly, BsmBI-v2.
New England Biolabs, Inc. Quick Blunting™ Kit – 20 reactions
The Quick Blunting Kit is used to convert DNA with incompatible 5' or 3' overhangs to 5' phosphorylated, blunt-ended DNA for efficient blunt-end ligation into DNA cloning vectors. DNA is blunted using T4 DNA Polymerase which has both 3' to 5' exonuclease activity and 5' to 3 polymerase activity. T4 Polynucleotide Kinase is included in the enzyme mix for phosphorylation of the 5' ends of blunt-ended DNA for subsequent ligation into a cloning vector. This kit is optimized for blunting up to 5 µg of DNA in a single reaction.
New England Biolabs, Inc. Q5 Site-Directed Mutagenesis Kit (Without Competent Cells) – 10 reactions
The Q5 Site-Directed Mutagenesis Kit (Without Competent Cells) enables rapid, site-specific mutagenesis of double-stranded plasmid DNA in less than 2 hours). The kit utilizes the robust Q5 Hot Start High-Fidelity DNA Polymerase along with custom mutagenic primers to create insertions, deletions and substitutions in a wide variety of plasmids. After PCR, the amplified material is added directly to a unique Kinase-Ligase-DpnI (KLD) enzyme mix for rapid (5 minutes), room temperature circularization and template removal. Transformation into high-efficiency chemically-competent E. coli, not supplied, ensures robust results with plasmids up to at least 20 kb in length. Kit is also available with competent cells .
New England Biolabs, Inc. RecA – 200 µg
E. coli RecA is necessary for genetic recombination, reactions involving DNA repair and UV-induced mutagenesis. RecA promotes the autodigestion of the lexA repressor, umuD protein and lambda repressor. Cleavage of LexA derepresses more than 20 genes. In vitro studies indicate that in the presence of ATP, RecA promotes the strand exchange of single-strand DNA fragments with homologous duplex DNA. The reaction has three distinct steps: (i) RecA polymerizes on the single-strand DNA, (ii) the nucleoprotein filament binds the duplex DNA and searches for a homologous region, (iii) the strands are exchanged.
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