Shop All DNA⁄RNA Modifying Enzymes

Ambion™ RNase A, affinity purified, 1 mg/mL Invitrogen™

Ambion® RNase A is an endonuclease that specifically cleaves 3' of U and C residues. Affinity Purified RNase A is intended for critical applications when the absence of DNase and other nonspecific nuclease activities is essential. Supplied in one tube containing 1 mg (1 mg/mL). Note: This preparation contains RNase B, a carbohydrate isoform of RNase A. RNase A is rigorously tested for contaminating nonspecific endonuclease, exonuclease, and protease activity.

RNase T1 (1 U/μL) Invitrogen™

Optimized for researchers performing RNA structure, RNA sequencing, protein footprinting, and boundary experiments, Ambion® RNase T1, RNA-Grade cleaves 3' of single-stranded G residues and is tested for purity to ensure that no contaminating ribonuclease activities are present that could cleave at unanticipated sites. Supplied in one tube of 200 µL (1U/ µL). RNase T1 can be used to perform boundary experiments to define the minimal RNA sequence required for selectable activities such as protein binding or catalysis. In addition to applications for RNA structural analysis, RNA-Grade ribonucleases can be used to map protein binding sites on RNAs by comparing cleavage patterns in the presence and absence of an RNA binding protein.

Unit Definitions:
100 units is the amount of enzyme that yields an increase in absorption at 260 nm of 0.01428 units per min at room temperature using 60 µg/mL yeast total RNA as a substrate.

Accessory Products:
Other RNA-Grade ribonuclease available include RNase A (SKU# AM2274), which cleaves 3' of single-stranded C and U residues. Combinations of the single- and double-stranded specific ribonucleases can provide rapid analysis of the physical structure of an interesting RNA.

RNase Cocktail™ Enzyme Mix Invitrogen™

Ambion® RNase Cocktail™ is a mixture of two highly purified ribonucleases, RNase A (500 U⁄ml) and RNase T1 (20,000 U⁄ml) and is free of DNase and nicking activities. Use RNase Cocktail for all situations where it is desirable to degrade RNA, i.e. plasmid minipreps and ribonuclease protection assays. RNase Cocktail is supplied in 50% glycerol for maximum convenience. Digestion of RNA with RNase A alone leaves fragments of RNA which are large enough to be visible on agarose gels and precipitate in ethanol. RNase A cuts after C and U residues, and RNase T1 cuts after G residues. Consequently, the mixture of both enzymes results in a reduction in RNA fragment size over the use of either alone.

Quality Control
RNase Cocktail Enzyme Mix is rigorously tested for contaminating nonspecific endonuclease, exonuclease, and protease activity. Functionality is determined in a ribonuclease protection assay.

Unit Definitions:
RNase A: One unit of RNase A is the amount required to give an increase in absorption at 286 nm of 0.0146 absorbance units per minute in a 1 mL volume. Unit assay conditions: 100 mM Tris-acetate (pH 6.5), 1 mM EDTA and 1 mM cyclic 2', 3'-CMP.

RNase T1: 100 Units of RNase T1 is the amount of enzyme that yields an increase in absorption at 260 nm of 0.01428 units per min at room temperature using 60 µg⁄mL yeast total RNA as a substrate.

We also now offer Ambion® RNA-Grade ribonucleases A, V1, and T1 for use in RNA structure⁄function studies. For more information, see: AM2274 RNase A (1 µg⁄ml), AM2275 RNase V1 (0.1 U⁄µl), AM2283 RNase T1 (1 U⁄µl).

Topoisomerase I Invitrogen™

Topoisomerase I (DNA-relaxing enzyme) catalyzes the removal of superhelical turns from covalently closed DNA by a transient breakage and rejoining of phosphodiester bonds. Topoisomerase I is active in the presence of EDTA.

Applications: Relaxing positively and negatively supercoiled DNA (1). Producing DNA topoisomers (2).

Source: Purified from calf thymus.

Performance and Quality Testing: Endodeoxyribonuclease, 3´ and 5´ exodeoxyribonuclease, and phosphatase assays; conversion of super-coiled DNA to relaxed DNA.

Unit Definition: One unit catalyzes the conversion of 0.5 µg of superhelical Φ X174 RF DNA to a relaxed state in 30 min. at 37°C.

Unit Reaction Conditions: 50 mM Tris-HCl (pH 7.5), 50 mM KCl, 10 mM MgCl2 , 0.1 mM EDTA, 0.5 mM DTT, 30 µg/ml/BSA, 0.5 µg
Φ X174 RF DNA, and enzyme in 50 µl for 30 min. at 37°C.

Ambion™ RNase III Invitrogen™

Ambion® Escherichia coli ribonuclease III (RNase III; EC 3.1.24) is a double-stranded RNA (dsRNA) specific endoribonuclease. In E. coli, RNase III cleaves ribosomal RNA (rRNA) precursors during maturation of rRNA. The enzyme cleaves dsRNA into 12–15 bp dsRNA fragments with 2 to 3 nucleotide 3' overhangs, and 5' phosphate and 3' hydroxyl termini. The termini and overhangs of RNase III cleavage products are thus the same as those produced by Dicer in the eukaryotic RNAi pathway. Transfection of RNase III cleavage products can be used to induce RNAi in mammalian cells (patent pending).

Unit Definition:
One unit is defined as the amount of enzyme catalyzing the cleavage of 1 µg of 500 bp dsRNA substrate to approximately 12–30 bp fragments in 60 min at 37°C.

T4 DNA Ligase (1 U/µL) Invitrogen™

T4 DNA Ligase catalyzes the formation of phosphodiester bonds in the presence of ATP between double-stranded DNAs with 3´ hydroxyl and 5´ phosphate termini. The unique T4 DNA Ligase buffer optimizes ligation, which can be performed in 5 minutes. Single-stranded nucleic acids are not substrates for this enzyme.

Applications
Cloning (blunt-end or cohesive-end ligation) and adding linkers or adapters to blunt-ended DNA

Source
Purified from E. coli lambda lysogen NM989

Performance and quality testing
Endodeoxyribonuclease, 3´ and 5´ exodeoxyribonuclease assays; ligation efficiency tested

Unit definition
One unit catalyzes the exchange of 1 nmol 32P-labeled pyrophosphate into ATP in 20 min at 37°C. One unit is equal to approximately 300 cohesive-end ligation units.

Unit reaction conditions
66 mM Tris-HCl (pH 7.6), 6.6 mM MgCl2, 10 mM DTT, 66 µM ATP, 3.3 µM 32P-labeled pyrophosphate, and enzyme in 0.1 mL for 20 min at 37°C.

Terminal Deoxynucleotidyl Transferase (20 U/µL) Thermo Scientific™

Thermo Scientific Terminal Deoxynucleotidyl Transferase (TdT) is a template-independent DNA polymerase that catalyzes the repetitive addition of deoxyribonucleotides to the 3'-OH of oligodeoxyribonucleotides and single-stranded and double-stranded DNA. TdT requires an oligonucleotide of at least three nucleotides to serve as a primer. With RNA as template TdT shows variable performance which strongly depends upon the tertiary structure of acceptor RNA 3'-end and the nature of nucleotide. Generally, it is lower than using DNA as a template.

Highlights

Incorporates modified nucleotides (e.g., fluorescein-, biotin-, aminoallyl-labeled nucleotides)

Applications

• Production of synthetic homo- and heteropolymers
• Homopolymeric tailing of linear duplex DNA with any type of 3'-OH terminus
• Oligodeoxyribonucleotide and DNA labeling
• 5'-RACE (Rapid Amplification of cDNA Ends)
In situ localization of apoptosis

Note

Due to the presence of CoCl2, the TdT Reaction Buffer is incompatible with downstream applications. It is necessary to remove CoCl2 from the reaction mixture by spin column or phenol/chloroform extraction and subsequent ethanol precipitation.

Endonuclease IV, E.coli (2 U/µL) Thermo Scientific™

Thermo Scientific Endonuclease IV (Endo IV) recognizes apurinic/apyrimidinic (AP) sites of dsDNA and cleaves the phosphodiester bond 5' to the lesion generating a hydroxyl group at the 3'-terminus (see Figure 1 in Supporting Data). The enzyme can also act as a 3'-diesterase that is able to release 3'-phosphoglycolate or 3'-phosphate from the damaged ends of dsDNA. Endo IV possesses also a 3'→5' exonuclease activity. Its progression on substrates is sensitive to ionic strength, metal ions, EDTA, and reducing conditions. Substrates with 3'-recessed ends are preferred substrates for the 3'→5' exonuclease activity.

The enzyme has no requirement for Mg2+ and is fully active in the presence of EDTA in moderate concentrations.

Applications

• Studies of DNA damage and repair
• Single cell electrophoresis (comet assay)
• Antitumor drug research
• DNA structure research
• SNP analysis

Ribonuclease H, from E. coli (cloned) 10 U/μL Invitrogen™

Ambion® RNase H (Ribonuclease H) is isolated from an E. coli strain that over-expresses the gene. RNase H specifically degrades the RNA in RNA:DNA hybrids to produce 3' -hydroxyl and 5' -phosphate terminated products. It is supplied in one tube containing 1,000 U (10 U/ µL). The enzyme will not degrade DNA or unhybridized RNA. RNase H is an integral part of most RNA amplification and NASBA protocols. It can also be used to degrade specific RNAs when the complementary DNA oligo is hybridized, such as poly(A) tail removal from mRNA hybridized to oligo(dT). Ribonuclease H is rigorously tested for contaminating nonspecific endonuclease, exonuclease, RNase, and protease activity.

Unit Definition:
One unit of Ribonuclease H is the amount of enzyme required to increase fluorescence 1.5 RFUs per sec at 37°C using 20 pmol of RNaseAlert® probe coupled to 1,000 pmol of a complementary oligonucleotide as substrate.

Terminal Deoxynucleotidyl Transferase, recombinant Invitrogen™

Terminal Deoxynucleotidyl Transferase, Recombinant (rTdT) is a DNA polymerase that catalyzes the addition of deoxynucleotides to the 3fi hydroxyl terminus of DNA. A TdT Technical Bulletin is available.

Applications:
Homopolymer tailing of vector and insert for cloning. Labeling oligonucleotides with biotin (1,2), 32P- or 35S-label (3), or in apoptosis (TUNEL) (4,5).

Source:
Purified from E. coli clone of calf thymus TdT.

Performance and Quality Testing:
Endonuclease, 3´ and 5´ exodeoxyribonuclease, and levels of incorporation tested.

Unit Definition:
One unit incorporates 1 nmol dATP into acid-precipitable material in 1 h at 37°C, using d(pA)50 as a primer.

Hazard Warning:
Toxic; potassium cacodylate contained in reaction buffer. Also contains cobalt chloride, a highly toxic chemical.
See MSDS.

Unit Reaction Conditions:
0.2 M potassium cacodylate (pH 7.2), 10 mM MgO4 C4 H6 , 1 mM 2-mercaptoethanol, 0.5 mg/ml BSA,
100 flM d(pA)50, 1 mM [3H]dATP, and enzyme in 0.15 ml for 1 h at 37°C.

T4 DNA Ligase, LC (1 U/µL) Thermo Scientific™

Thermo Scientific T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA. The enzyme repairs single-strand nicks in duplex DNA, RNA, or DNA/RNA hybrids. It also joins DNA fragments with either cohesive or blunt termini, but has no activity on single-stranded nucleic acids.

T4 DNA Ligase requires ATP as a cofactor.

Highlights

• Active in Themo Scientific restriction enzyme, PCR, and RT buffers (when supplemented with ATP)
• Fast—sticky-end ligation is completed in 10 minutes at room temperature
• Supplied with PEG solution for efficient blunt-end ligation

Applications

• Cloning of restriction enzyme generated DNA fragments
• Cloning of PCR products
• Joining of double-stranded oligonucleotide linkers or adaptors to DNA
• Site-directed mutagenesis
• Amplified fragment length polymorphism (AFLP)
• Ligase-mediated RNA detection (see Reference 3)
• Nick repair in duplex DNA, RNA or DNA/RNA hybrids
• Self-circularization of linear DNA.

Includes

• T4 DNA Ligase
• 10X T4 DNA Ligase Buffer
• 50% PEG Solution

Notes

• Binding of T4 DNA Ligase to DNA may result in a band shift in agarose gels. To avoid this, incubate samples with 6X DNA Loading Dye & SDS Solution at 70°C for 5 min or 65°C for 10 minutes and chill on ice prior to electrophoresis.
• The volume of the ligation reaction mixture should not exceed 10% of the competent cell volume in the transformation process.
• Prior to electro-transformation, remove T4 DNA Ligase from the ligation mixture using spin columns or chloroform extraction. The extracted DNA can be further precipitated with ethanol.

DNase I, Amplification Grade Invitrogen™

DNase I, Amplification Grade, digests single- and double-stranded DNA to oligodexyribonuleotides containing a 5' phosphate. DNase I, Amplification Grade, is suitable for eliminating DNA during critical RNA purification procedures such as those prior to RNA-PCR amplification. It is purified and tested for non-detectable levels of RNase contamination. Absence of RNase is tested by performing a ribonuclease assay with RNA ladder.

Application
Removing DNA from RNA and protein preparations.

Specific activity
Specific activity is >10,000 units/mg.

Source
Purified from bovine pancreas

Performance and quality testing
Ribonuclease assay with RNA ladder and ability to digest single-stranded and double-stranded DNA to oligonucleotides are determined.

Unit definition
One unit increases the absorbance of a high molecular weight DNA solution at a rate of 0.001 A260 units/min/mL of reaction mixture at 25°C.

Unit reaction conditions
0.1 M sodium acetate (pH 5.0), 5 mM MgCl2, 50 µg/mL calf thymus DNA, and enzyme in 1 mL for 10 min at 25°C.

RNase H (5 U/µL) Thermo Scientific™

Thermo Scientific Ribonuclease H (RNase H) specifically degrades the RNA strand in RNA-DNA hybrids. It does not hydrolyze the phosphodiester bonds within single-stranded and double-stranded DNA and RNA.

Applications

• Removal of mRNA prior to synthesis of second strand cDNA
RT-PCR and qRT-PCR: removal of RNA after first strand cDNA synthesis
• Removal of the poly(A) sequences of mRNA after hybridization with oligo(dT)
• Site-specific cleavage of RNA
• Studies of in vitro polyadenylation reaction products

T4 DNA Ligase (5 U/µL) Thermo Scientific™

Thermo Scientific T4 DNA Ligase catalyzes the formation of a phosphodiester bond between juxtaposed 5'-phosphate and 3'-hydroxyl termini in duplex DNA or RNA. The enzyme repairs single-strand nicks in duplex DNA, RNA, or DNA/RNA hybrids. It also joins DNA fragments with either cohesive or blunt termini, but has no activity on single-stranded nucleic acids.

T4 DNA Ligase requires ATP as a cofactor.

Highlights

• Active in Themo Scientific restriction enzyme, PCR, and RT buffers (when supplemented with ATP)
• Fast—sticky-end ligation is completed in 10 minutes at room temperature
• Supplied with PEG solution for efficient blunt-end ligation

Applications

• Cloning of restriction enzyme generated DNA fragments
• Cloning of PCR products
• Joining of double-stranded oligonucleotide linkers or adaptors to DNA
• Site-directed mutagenesis
• Amplified fragment length polymorphism (AFLP)
• Ligase-mediated RNA detection (see Reference 3)
• Nick repair in duplex DNA, RNA or DNA/RNA hybrids
• Self-circularization of linear DNA.

Includes

• T4 DNA Ligase
• 10X T4 DNA Ligase Buffer
• 50% PEG Solution

Notes

• Binding of T4 DNA Ligase to DNA may result in a band shift in agarose gels. To avoid this, incubate samples with 6X DNA Loading Dye & SDS Solution at 70°C for 5 min or 65°C for 10 minutes and chill on ice prior to electrophoresis.
• The volume of the ligation reaction mixture should not exceed 10% of the competent cell volume in the transformation process.
• Prior to electro-transformation, remove T4 DNA Ligase from the ligation mixture using spin columns or chloroform extraction. The extracted DNA can be further precipitated with ethanol.

T4 beta-glucosyltransferase Thermo Scientific™

Thermo Scientific T4 β-glucosyltransferase (T4 BGT) transfers the glucose moiety of uridine diphosphoglucose (UDP-glucose) to the 5-hydroxymethylcytosine (5-hmC) residues in double-stranded DNA generating ß-glucosyl-5 hydroxymethylcytosine.

Thermo Scientific T4 BGT is specifically formulated for fast reaction times without compromising the reaction efficiency. The enzyme completes 5-hmC glucosylation of 1 µg DNA at 37°C in 15 minutes.

Highlights

Specific—selectively transfers glucose to the hydroxymethyl moiety of 5-hmC
Fast—complete glucosylation of 1 µg DNA in just 15 min
Convenient—supplied with optimized buffer and UDP-glucose.

Applications

• Locus specific detection of 5-hmC
• Enrichment of 5-hmC containing DNA
• Labeling of 5-hmC residues using a radioactive UDP-glucose donor

Includes
• T4 β-glucosyltransferase, 5 U/µL
• 10X Epi buffer
• 10X UDP-glucose
Results per page
    spinner