Shop All PCR & Cloning Enzymes

S1 Nuclease (Invitrogen™)

S1 Nuclease is a single-strand-specific endonuclease that hydrolyzes single-stranded RNA or DNA into 5´ mononucleotides. The enzyme will hydrolyze single-stranded regions in duplex DNA such as loops and gaps. S1 Nuclease is stable at 65°C.

Applications:
Nuclease mapping techniques (1,2). Removal of single-stranded regions from double-stranded DNA (3). Exo III-ordered sequencing (4).

Source:
Isolated from Aspergillus oryzae.

Performance and Quality Testing:
Double-strand-specific deoxyribonuclease and phosphatase assays.

Unit Definition:
One unit hydrolyzes 1 µg of denatured DNA to acid-soluble material in 1 min. at 37°C.

Unit Reaction Conditions:
30 mM sodium acetate (pH 4.6), 50 mM NaCl, 1 mM zinc acetate, 0.5 mg/ml heat-denatured DNA, 5% (v/v) glycerol, and enzyme in 0.5 ml for 10 min. at 37°C.

RNase T1 (1000 U/µL) (Thermo Scientific™)

Thermo Scientific RNase T1 is an endoribonuclease that specifically degrades single-stranded RNA at G residues. It cleaves the phosphodiester bond between the 3'-guanylic residue and the 5'-OH residue of adjacent nucleotides with the formation of corresponding intermediate 2', 3'-cyclic phosphate. The reaction products are 3'-GMP and oligonucleotides with a terminal 3'-GMP.

RNase T1 does not require metal ions for activity.

Applications

• Removal of RNA from DNA preparations
• RNA sequencing
• Ribonuclease protection assays. Used in conjunction with RNase A
• Removal of RNA from recombinant protein preparations
• Determination of the level of RNA transcripts synthesized in vitro from DNA templates containing a "G-less cassette"

TaqI (10 U/µL) (Thermo Scientific™)

5'  T ↓C  G  A   3' 
3'  A  G  C ↑T   5' 

Thermo Scientific TaqI restriction enzyme recognizes T^CGA sites and cuts best at 65°C in its own unique buffer. See Reaction Conditions for Restriction Enzymes for a table of enzyme activity, conditions for double digestion, and heat inactivation for this and other restriction enzymes. Note: Also available as a FastDigest enzyme for rapid DNA digestion.

Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.

Features

• Superior quality—stringent quality control and industry leading manufacturing process
• Convenient color-coded Five Buffer System
• Includes universal Tango buffer for double-digestions
• BSA premixed in reaction buffers
• Wide selection of restriction endonuclease specificities

Applications

• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern blotting
• Restriction fragment length polymorphism (RFLP)
• SNP

Note: Assayed using Lambda DNA (dam-) (#SD0021). TaqI is blocked by overlapping dam methylation. To avoid dam methylation, use a dam-, dcm- strain, such as GM2163. Incubation at 37°C results in 10% activity. We recommend using TaqI, HC at 37°C. For methylation sensitivity, refer to product specifications.

FastDigest PvuI (Thermo Scientific™)

5'  C  G  A  T ↓C  G   3' 
3'  G  C ↑T  A  G  C   5' 

Thermo Scientific FastDigest PvuI restriction enzyme recognizes CGAT^CG site and cuts best at 37°C in 5–15 minutes using universal FastDigest Buffer. Isoschizomers: BpvUI, MvrI, Ple19I.

Thermo Scientific FastDigest PvuI is one of an advanced line of fast restriction enzymes that are all 100% active in the universal FastDigest and FastDigest Green reaction buffers.

The universal buffer allows rapid single-, double-, or multiple DNA digestion within 5–15 minutes eliminating any need for buffer change or subsequent DNA clean-up steps. See Reaction Conditions for FastDigest Enzymes for a table of enzyme activity, heat inactivation and incubation times for this and other FastDigest restriction enzymes. DNA modifying enzymes, such as Klenow Fragment, T4 DNA Ligase, alkaline phosphatases and T4 DNA Polymerase all have 100% activity in FastDigest Buffer. Therefore, enzymes for downstream applications can be directly added to the FastDigest reaction mix.

For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels.

Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects.

Features

• 100% activity of all FastDigest enzymes in the universal buffer
• 100% buffer compatibility with downstream applications
• Complete digestion in 5–15 minutes
• Direct loading on gels
• No star activity
• 176 FastDigest enzymes available

Applications

• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern Blot
• Restriction fragment length polymorphism (RFLP)
• SNP analysis

Note: The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer. For applications that require product analysis by fluorescence excitation (e.g. concentration measurements in UV light) the colorless FastDigest Buffer is recommended.

For methylation sensitivity, refer to product specifications.

DNA Polymerase I, Large (Klenow) Fragment (Invitrogen™)

DNA Polymerase I, Large (Klenow) Fragment is a DNA polymerase enzyme that lacks the 5' to 3' exonuclease activity of intact DNA Polymerase I, but does exhibit the 5' to 3' DNA polymerase and 3' to 5' exonuclease activities.

Applications:
Fill-in of 5´ overhangs (1). Synthesis of probes by random primers labeling method (2). Sequencing single- and double-stranded DNA (3). Site-directed mutagenesis.

Source:
Purified from E. coli expressing the Klenow fragment on a plasmid.

Performance and Quality Testing:
SDS-PAGE purity; single-stranded and double-stranded endodeoxyribonuclease and self-priming assays; performance evaluated in fill-in reaction.

Unit Definition:
One unit incorporates 10 nmol of total deoxyribonucleotide into acid-precipitable material in 30 min. at 37°C using a template•primer.

AccuPrime™ GC-Rich DNA Polymerase (Invitrogen™)

AccuPrime™ GC-Rich DNA Polymerase is designed to provide high-yield, high-specificity amplification of difficult-to-amplify templates, such as those with >65% GC content. The kit offers a choice of buffers for amplifying genomic DNA targets (Buffer A) or non-GC-rich cDNA, plasmid, and lambda-based targets (Buffer B). Benefits of using AccuPrime™ GC-Rich DNA Polymerase:

Yield—high yields for targets up to 5 kb in length
Specificity—AccuPrime™ accessory proteins for improved PCR specificity
Sensitivity—sensitivity down to 5 ng of template DNA

Robust and specific amplifications
The AccuPrime™ buffers contain thermostable proteins that enhance primer-template hybridization during PCR, increasing the specificity of the reaction. The polymerase itself, from the archaebacterium Pyrolobus fumarius, has a five-fold better processivity than Taq DNA polymerase, and remains active even after 4 hours at 95°C.

Unit definition
One unit of AccuPrime™ GC-Rich DNA Polymerase is the amount of enzyme required to incorporate 10 nmoles of dNTPs into acid-insoluble material in 30 min at 74°C.

Anza™ 102 CaiI (Invitrogen™)

5'  C  A  G  N  N  N ↓C  T  G   3' 
3'  G  T  C ↑N  N  N  G  A  C   5' 

Invitrogen™ Anza™ 102 CaiI is a restriction enzyme that cuts DNA at this recognition site: CAGNNN^CTG, completely digesting the DNA in 15 minutes at 37°C. Isoschizomers include: AlwNI, PstNI.

For superior convenience, a single buffer and protocol are used with all Anza™ restriction enzymes. The single Anza buffer allows multiple restriction enzymes to be used simultaneously in one reaction. This helps eliminate the need for sequential digests and the DNA clean-up necessary when changing buffers. Additionally, the Anza buffer and restriction enzymes have been designed for flexibility in digestion times. Digestions of up to 16 hours can be carried out without star activity.

Convenient—all Anza enzymes are 100% functional in the single Anza buffer
Fast—complete digestion in 15 minutes
Flexible—overnight digestion without star activity
Simple—a single digestion protocol for all DNA types
Complete system—incorporates Anza DNA Modifying Enzymes for a complete cloning experience

Anza DNA Modifying Enzymes include the Anza T4 DNA Ligase Master Mix, Anza Alkaline Phosphatase Kit, Anza T4 PNK Kit, Anza DNA Blunt End Kit, and Anza DNA End Repair Kit. Use these enzymes for a complete cloning workflow.

Anza Red Buffer, which contains a density gradient and tracking dyes, is supplied for added convenience. Digestion reactions are as efficient in the Anza Red Buffer as in the Anza Buffer. Use of the Anza Red Buffer helps reduce pipetting steps when preparing multiple digestion reactions and a subsequent step includes agarose gel electrophoresis. For applications that require product analysis by fluorescence excitation, the colorless Anza Buffer without the tracking dyes is recommended.

Anza™ 41 HpyF3I (Invitrogen™)

5'  C ↓T  N  A  G   3' 
3'  G  A  N  T ↑C   5' 

Invitrogen™ Anza™ 41 HpyF3I is a restriction enzyme that cuts DNA at this recognition site: C^TNAG, completely digesting the DNA in 15 minutes at 37°C. Isoschizomers include: BstDEI, DdeI.

For superior convenience, a single buffer and protocol are used with all Anza™ restriction enzymes. The single Anza buffer allows multiple restriction enzymes to be used simultaneously in one reaction. This helps eliminate the need for sequential digests and the DNA clean-up necessary when changing buffers. Additionally, the Anza buffer and restriction enzymes have been designed for flexibility in digestion times. Digestions of up to 16 hours can be carried out without star activity.

Convenient—all Anza enzymes are 100% functional in the single Anza buffer
Fast—complete digestion in 15 minutes
Flexible—overnight digestion without star activity
Simple—a single digestion protocol for all DNA types
Complete system—incorporates Anza DNA Modifying Enzymes for a complete cloning experience

Anza DNA Modifying Enzymes include the Anza T4 DNA Ligase Master Mix, Anza Alkaline Phosphatase Kit, Anza T4 PNK Kit, Anza DNA Blunt End Kit, and Anza DNA End Repair Kit. Use these enzymes for a complete cloning workflow.

Anza Red Buffer, which contains a density gradient and tracking dyes, is supplied for added convenience. Digestion reactions are as efficient in the Anza Red Buffer as in the Anza Buffer. Use of the Anza Red Buffer helps reduce pipetting steps when preparing multiple digestion reactions and a subsequent step includes agarose gel electrophoresis. For applications that require product analysis by fluorescence excitation, the colorless Anza Buffer without the tracking dyes is recommended.

AmpliTaq™ DNA Polymerase, LD (Low DNA) with Buffer II (Applied Biosystems™)

Applied Biosystems® AmpliTaq® DNA Polymerase, LD (Low DNA), is the same enzyme as AmpliTaq® DNA Polymerase, but is further purified through a proprietary separation process to ensure that residual bacterial DNA sequences are substantially reduced.

Manufacturing process results in highly purified enzyme with lower levels of bacterial DNA

• Minimizes non-specific PCR products when amplifying bacterial targets
• Highly purified enzyme making it particularly useful for low copy number amplifications

High-Quality
This highly purified enzyme preparation ensures that non-target, false-positive PCR products will be effectively minimized when amplifying bacterial sequences. This is especially useful for low-copy-number PCR amplifications. The enzyme is quality-control tested to verify that fewer than 10 copies of bacterial 16S ribosomal RNA gene sequences are present in a standard 2.5 unit aliquot. AmpliTaq® DNA Polymerase, LD is supplied with GeneAmp® 10X PCR Buffer II and MgCl2 Solution. It is also available with GeneAmp® 10X PCR Buffer I.

Note:
See user's manual or package insert for limited label license and trademark information. For Research Use Only. Not for human or animal therapeutic or diagnostic use.

DreamTaq DNA Polymerase (5 U/µL) (Thermo Scientific™)

Thermo Scientific DreamTaq DNA Polymerase is an enhanced Taq DNA polymerase optimized for all standard PCR applications. It ensures higher sensitivity, longer PCR products and higher yields compared to conventional Taq DNA polymerase. DreamTaq DNA Polymerase uses the same reaction set-up and cycling conditions as conventional Taq DNA polymerase. Extensive optimization of reaction conditions is not required. It is supplied with optimized DreamTaq buffer, which includes 20 mM MgCl2.

Highlights

• Robust amplification with minimal optimization
• High yields of PCR products
• Higher sensitivity compared to conventional Taq DNA polymerase
• Amplification of long targets up to 6 kb from genomic DNA and up to 20 kb from viral DNA
• Incorporates modified nucleotides, but does not incorporate dUTP

Applications

• Routine PCR amplification of DNA fragments up to 6 kb
• High throughput PCR
• Genotyping

Anza™ 9 NdeI (Invitrogen™)

5'  C  A ↓T  A  T  G   3' 
3'  G  T  A  T ↑A  C   5' 

Invitrogen™ Anza™ 9 NdeI is a restriction enzyme that cuts DNA at this recognition site: CA^TATG, completely digesting the DNA in 15 minutes at 37°C. Isoschizomers include: FauNDI.

For superior convenience, a single buffer and protocol are used with all Anza™ restriction enzymes. The single Anza buffer allows multiple restriction enzymes to be used simultaneously in one reaction. This helps eliminate the need for sequential digests and the DNA clean-up necessary when changing buffers. Additionally, the Anza buffer and restriction enzymes have been designed for flexibility in digestion times. Digestions of up to 16 hours can be carried out without star activity.

Convenient—all Anza enzymes are 100% functional in the single Anza buffer
Fast—complete digestion in 15 minutes
Flexible—overnight digestion without star activity
Simple—a single digestion protocol for all DNA types
Complete system—incorporates Anza DNA Modifying Enzymes for a complete cloning experience

Anza DNA Modifying Enzymes include the Anza T4 DNA Ligase Master Mix, Anza Alkaline Phosphatase Kit, Anza T4 PNK Kit, Anza DNA Blunt End Kit, and Anza DNA End Repair Kit. Use these enzymes for a complete cloning workflow.

Anza Red Buffer, which contains a density gradient and tracking dyes, is supplied for added convenience. Digestion reactions are as efficient in the Anza Red Buffer as in the Anza Buffer. Use of the Anza Red Buffer helps reduce pipetting steps when preparing multiple digestion reactions and a subsequent step includes agarose gel electrophoresis. For applications that require product analysis by fluorescence excitation, the colorless Anza Buffer without the tracking dyes is recommended.

Phusion Green Hot Start II High-Fidelity DNA Polymerase (2 U/µL) (Thermo Scientific™)

Thermo Scientific Phusion High-Fidelity DNA Polymerases set a gold standard for high performance PCR. Featuring an error rate 50-fold lower than that of Taq, and 6-fold lower than that of Pfu, Phusion High-Fidelity DNA Polymerase is excellent choice for cloning and other applications requiring high fidelity. Phusion DNA Polymerases offer robust performance with short protocol times, even in the presence of PCR inhibitors, and generate higher yields with lower enzyme amounts than other DNA polymerase.

With Phusion Hot Start II High-Fidelity DNA Polymerase amplification proceeds without the production of nonspecific products due to the combination of Phusion DNA Polymerase and a reversibly bound, specific Affibody ligand that inhibits DNA polymerase activity at room temperature. The Affibody ligand also inhibits the 3´→5´ exonuclease activity of the polymerase, thus preventing degradation of primers and template DNA during reaction set up. At temperatures that promote polymerase activity, the ligand is released, rendering the polymerase fully active. Phusion Hot Start II DNA Polymerase is immediately reactivated at high temperatures so it does not require a separate activation step in PCR protocols.
Phusion Green Hot Start II High-Fidelity DNA Polymerase is a combination of Phusion Hot Start II DNA Polymerase and the 5X Phusion Green Buffers. The buffers include a density reagent and two tracking dyes which do not interfere with the robust performance of Phusion Hot Start II DNA Polymerase. Researchers may directly load PCR products on a gel making this a versatile option for use in downstream applications such as DNA sequencing, ligation and restriction digestion.

Highlights

• Reaction set up at room temperature
• No non-specific amplification and primer degradation during reaction set up
• Zero-time reactivation due to unique hot start technology
• High fidelity (52X Taq)
• Fast PCR due to short extension times (15-30 s/kb)
• Robust reactions, minimal optimization needed
• Increased product yields with minimal enzyme amounts
• Direct loading on gels

Applications

• High-fidelity PCR
• High throughput
• Amplification of difficult (GC-rich) templates
• Template generation for sequencing
• Multiplex PCR
• Long-range PCR
• Cloning
• Mutagenesis
• Microarray

Using Phusion DNA Polymerases
Annealing rules for Phusion DNA Polymerases are different from many common DNA polymerases (such as Taq DNA polymerases). For optimal results, use our Tm calculator at www.thermofisher.com/tmcalculator.

FastDigest PvuII (Thermo Scientific™)

5'  C  A  G ↓C  T  G   3' 
3'  G  T  C ↑G  A  C   5' 

Thermo Scientific FastDigest PvuII restriction enzyme recognizes CAG^CTG site and cuts best at 37°C in 5–15 minutes using universal FastDigest Buffer.

Thermo Scientific FastDigest PvuII is one of an advanced line of fast restriction enzymes that are all 100% active in the universal FastDigest and FastDigest Green reaction buffers.

The universal buffer allows rapid single-, double-, or multiple DNA digestion within 5–15 minutes eliminating any need for buffer change or subsequent DNA clean-up steps. See Reaction Conditions for FastDigest Enzymes for a table of enzyme activity, heat inactivation and incubation times for this and other FastDigest restriction enzymes. DNA modifying enzymes, such as Klenow Fragment, T4 DNA Ligase, alkaline phosphatases and T4 DNA Polymerase all have 100% activity in FastDigest Buffer. Therefore, enzymes for downstream applications can be directly added to the FastDigest reaction mix.

For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels.

Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects.

Features

• 100% activity of all FastDigest enzymes in the universal buffer
• 100% buffer compatibility with downstream applications
• Complete digestion in 5–15 minutes
• Direct loading on gels
• No star activity
• 176 FastDigest enzymes available

Applications

• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern Blot
• Restriction fragment length polymorphism (RFLP)
• SNP analysis

Note: The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer. For applications that require product analysis by fluorescence excitation (e.g. concentration measurements in UV light) the colorless FastDigest Buffer is recommended.

For methylation sensitivity, refer to product specifications.

FastDigest NdeI (Thermo Scientific™)

5'  C  A ↓T  A  T  G   3' 
3'  G  T  A  T ↑A  C   5' 

Thermo Scientific FastDigest NdeI restriction enzyme recognizes CA^TATG site and cuts best at 37°C in 5–15 minutes using universal FastDigest Buffer. Isoschizomers: FauNDI.

Thermo Scientific FastDigest NdeI is one of an advanced line of fast restriction enzymes that are all 100% active in the universal FastDigest and FastDigest Green reaction buffers.

The universal buffer allows rapid single-, double-, or multiple DNA digestion within 5–15 minutes eliminating any need for buffer change or subsequent DNA clean-up steps. See Reaction Conditions for FastDigest Enzymes for a table of enzyme activity, heat inactivation and incubation times for this and other FastDigest restriction enzymes. DNA modifying enzymes, such as Klenow Fragment, T4 DNA Ligase, alkaline phosphatases and T4 DNA Polymerase all have 100% activity in FastDigest Buffer. Therefore, enzymes for downstream applications can be directly added to the FastDigest reaction mix.

For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels.

Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects.

Features

• 100% activity of all FastDigest enzymes in the universal buffer
• 100% buffer compatibility with downstream applications
• Complete digestion in 5–15 minutes
• Direct loading on gels
• No star activity
• 176 FastDigest enzymes available

Applications

• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern Blot
• Restriction fragment length polymorphism (RFLP)
• SNP analysis

Note: The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer. For applications that require product analysis by fluorescence excitation (e.g. concentration measurements in UV light) the colorless FastDigest Buffer is recommended.

For methylation sensitivity, refer to product specifications.

MspI (HpaII) (10 U/µL) (Thermo Scientific™)

5'  C ↓C  G  G   3' 
3'  G  G  C ↑C   5' 

Thermo Scientific MspI (HpaII) restriction enzyme recognizes C^CGG sites and cuts best at 37°C in Tango buffer. See Reaction Conditions for Restriction Enzymes for a table of enzyme activity, conditions for double digestion, and heat inactivation for this and other restriction enzymes. Note: Also available as a FastDigest enzyme for rapid DNA digestion.

Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.

Features

• Superior quality—stringent quality control and industry leading manufacturing process
• Convenient color-coded Five Buffer System
• Includes universal Tango buffer for double-digestions
• BSA premixed in reaction buffers
• Wide selection of restriction endonuclease specificities

Applications

• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern Blot
• Restriction fragment length polymorphism (RFLP)
• SNP

Note: MspI is an isoschizomer of HpaII. When the external C in the sequence CCGG is methylated, MspI and HpaII cannot cleave. However, unlike HpaII, MspI can cleave the sequence when the internal C residue is methylated. For methylation sensitivity, refer to product specifications.