DNA Modifying Enzymes and Cloning Technologies

• Apurinic/apyrimidinic Endonuclease 1
• DNA AP endonuclease that catalyzes the cleavage of DNA phosphodiester backbone at AP sites via hydrolysis leaving a 1 nucleotide gap with 3′-hydroxyl and 5′ deoxyribose phosphate (dRP) termini
• APE 1 has also been reported to have DNA 3′-diesterase, 3′ to 5′ exonuclease, and RNase H activities
• Also known as HAP1 or Ref-1

Blunt/TA Ligase Master Mix is a ready-to-use solution of T4 DNA Ligase, proprietary ligation enhancer, and optimized reaction buffer. This master mix is specifically formulated to improve ligation and transformation of both blunt-end and single-base overhang substrates.

• Simplifies reaction set-up, ensures an optimized ratio of enzyme and buffer components
• No thawing is necessary as it remains liquid during storage at -20°C
• Ligations for subcloning can be carried out in small volumes with low DNA concentrations, allowing users to conserve precious DNA samples and directly transform many strains of chemically competent E. coli without dilution
• Not sure which ligase to choose? Refer to our DNA and RNA Ligase Properties Chart

• Ideal for in vitro digestion of dsDNA
• Compatible with EnGen^® sgRNA Synthesis Kit, S. pyogenes (NEB #E3322S) and the EnGen Mutation Detection Kit (NEB #E3321S)
• For help with oligo design, try EnGen sgRNA Template Oligo Designer – now included in the NEBioCalculator^® Tool

• Endonuclease that nonspecifically cleaves DNA to release di-, tri- and oligonucleotide products with 5′-phosphorylated and 3′-hydroxylated ends
• Degrades DNA template in transcription reactions
• Removes contaminating genomic DNA from RNA samples
• Isolated from a recombinant source

• Ideal for direct introduction of Cas9/sgRNA complexes
• Dual NLS for improved transport to the nucleus
• Compatible with EnGen^® sgRNA Synthesis Kit, S. pyogenes (NEB #E3322S) and the EnGen Mutation Detection Kit (NEB #E3321S)

• Variant of Cas9 nuclease differing by a point mutation (D10A) in the RuvC nuclease domain
• Capable of generating nicks, but not cleaving DNA
• DNA double strand breaks can be generated, with reduced off-target cleavage, by targeting two sites with EnGen Cas9 Nickase in close proximity

Characteristics of Hi-T4 DNA Ligase:

• Hi-T4 DNA Ligase is an engineered variant of T4 DNA Ligase with improved thermostability compared to T4 DNA Ligase
• Capable of joining blunt end and cohesive end termini, as well as repairing single stranded nicks in duplex DNA and some DNA/RNA hybrids
• Supplied with both T4 DNA Ligase Buffer and StickTogether^™ DNA Ligase Buffer
• Other T4 DNA Ligase products include Quick Ligation Kit, Salt-T4, Instant Sticky-end Ligase Master Mix and Blunt/TA Ligase Master Mix
• Not sure which ligase to choose? Refer to our DNA and RNA Ligase Properties Chart or DNA Ligase Selection Chart

The NEBNext® Microbiome DNA Enrichment Kit facilitates enrichment of microbial DNA from samples containing methylated host DNA (including human), by selective binding and removal of the CpG-methylated host DNA. Importantly, microbial diversity remains intact after enrichment.

• Effective enrichment of microbial genomic DNA from samples contaminating host DNA
• Fast, simple protocol
• Enables microbiome whole genome sequencing, even for samples with high levels of host DNA
• Compatible with downstream applications including next-generation sequencing on all platforms, qPCR and endpoint PCR
• Suitable for a wide range of sample types
• No requirement for live cells
• Optional protocol to retain separated host DNA
• Also effective for separation of organelle DNA (e.g. mitochondria, chloroplast) from eukaryote nuclear DNA

This module is part of the original standard library prep workflow, which is longer than the NEBNext Ultra II DNA Library Prep Kit (NEB #E7645) 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.

This module is part of the original standard library prep workflow, which is longer than the NEBNext® Ultra II DNA Library Prep Kit (NEB #E7645) 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.

This module is part of the original standard library prep workflow, which is longer than the NEBNext^® Ultra II DNA Library Prep Kit (NEB #E7645) 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.

This module is also compatible with some Oxford Nanopore Technologies^® workflows, including some involving COVID-19 research.

Quick CIP from NEB is a heat-labile version of calf intestinal alkaline phosphatase (CIP) purified from a recombinant source.

Features of Quick CIP, NEB:

• Rapid and irreversible heat inactivation eliminates the unwanted activity
• Improved storage stability versus native enzyme
• Faster reaction setup (no supplemental additives like zinc required) and shorter incubation time
• Flexible reaction conditions (active in any restriction enzyme buffer, no clean-up required)
• Less enzyme required (high specific activity), resulting in a lower cost per reaction
• No need for multiple phosphatases (Quick CIP removes 5′- and 3′- phosphates from DNA, RNA and dNTPs )
• Active on unincorporated dNTPs in PCR products – improve DNA sequencing and SNP analysis
• Recombinant for purity, consistency and value

The Quick Ligation™ Kit enables ligation of cohesive end or blunt end DNA fragments in 5 minutes at room temperature. (25°C )

• Fast – 5 minutes for cohesive or blunt ends
• Convenient – ligation performed at room temperature
• Flexible – suitable for all common ligation reactions
• Not sure which ligase to choose? Refer to our DNA and RNA Ligase Properties Chart

Characteristics of Salt-T4 DNA Ligase:

• Salt-T4 DNA Ligase is an engineered variant of T4 DNA Ligase with improved salt tolerance
• Capable of joining blunt end and cohesive end termini, as well as repairing single stranded nicks in duplex DNA and some DNA/RNA hybrids.
• Supplied with both T4 DNA Ligase Buffer and StickTogether™ DNA Ligase Buffer.
• Other T4 DNA Ligase products include Quick Ligation Kit, Hi-T4, Instant Sticky-end Ligase Master Mix and Blunt/TA Ligase Master Mix
• Not sure which ligase to choose? Refer to our DNA and RNA Ligase Properties Chart or DNA Ligase Selection Chart

Shrimp Alkaline Phosphatase (rSAP) is a heat labile alkaline phosphatase purified from a recombinant source.

• Rapid and irreversible heat inactivation eliminates unwanted activity
• Improved storage stability versus native enzyme
• Faster reaction setup (no supplemental additives like zinc required)
• Flexible reaction conditions (active in any restriction enzyme buffer, no clean-up required)
• Less enzyme required (high specific activity), resulting in a lower cost per reaction
• No need for multiple phosphatases (rSAP removes 5´- and 3´- phosphates from DNA, RNA and dNTPs )
• Active on unincorporated dNTPs in PCR products – improves DNA sequencing and SNP analysis
• Recombinant for purity, consistency and value

Characteristics of T4 DNA Ligase, NEB:

• Catalyzes the formation of a phosphodiester bond between juxtaposed 5′ phosphate and 3′ hydroxyl termini in duplex DNA or RNA.
• This enzyme will join blunt end and cohesive end termini as well as repair single-stranded nicks in duplex DNA and some DNA/RNA hybrids (1).
• T4 DNA ligase will seal nicks for these DNA substrates.
• The industry standard for performance and quality
• Not sure which ligase to choose? Refer to our DNA and RNA Ligase Properties Chart