Thermostable RNase H
Cat. No.: SB-G3418-250U
Size: 250U
Description
This product, RNase H (Thermostable), is heat-resistant ribonuclease H, derived from Thermus thermophilus , and is an endoribonuclease that remains active at relatively high temperatures (above 65°C). RNase H (Thermostable) has similar enzymatic properties to the common E.coli RNase H, which specifically hydrolyzes the RNA in the DNA-RNA hybrid strand, and the hydrolysis products are oligoribonucleotides and single-strands at the 5' phosphate end DNA, cannot hydrolyze single- or double-stranded DNA or RNA. RNase H (Thermostable) still has high enzymatic activity above 65°C, its half-life can reach several hours at 70°C, and about 30 minutes at 95°C. This product is similar to the common E.coli RNase H. It is mainly used to remove mRNA before the second strand of cDNA is synthesized, remove poly(A) in mRNA hybridized with poly(dT), and realize the fixed point of RNA through DNA sequence cut etc.
- Source : derived from thermophilic bacteria, recombinantly expressed by Escherichia coli.
- Definition of enzyme activity : at 50°C, in 50 μL system, hydrolyze 40 pmol fluorescently labeled hybrid strand (RNA/DNA) substrate containing 25 base pairs within 20 minutes to produce 1 nmol ribonucleotide The required amount of enzyme is defined as one enzyme activity unit.
- Purity and concentration : SDS-PAGE detection purity ≥ 95%; 5 U/μL.
- Inactivation or inhibition :Add appropriate amount of proteinase K or add 5% volume of 0.5 M EDTA.
- Enzyme storage buffer : 50 mM Tris-HCl, 1 M NaCl, 0.1 mM EDTA, 1 mM DTT, 50% Glycerol, pH 7.5.
- 10 × Reaction Buffer : 500 mM Tris-HCl, 30 mM MgCl2 , 100 mM DTT, pH 8.3.
Storage and Handling Conditions
Transport in wet ice packs; store at -20°C, valid for 12 months.
Composition
| Component Number |
Component |
G3418-250U |
| G3418-1 |
Thermostable RNase H |
50 μL |
| G3418-2 |
10 × Reaction Buffers |
200 μL |
| Product Manual |
1 copy |
Steps
Removal of RNA in DNA-RNA hybrids:
1. Configure the reaction system according to the following table:
| Component |
Volume |
| Thermostable RNase H |
1 μL |
| 10 × Reaction Buffers |
2 μL |
| DNA-RNA hybrid strand |
≤2μg |
| Nuclease Free Water |
To 20 μL |
Note: The DNA-RNA hybrid strand in the above system does not exceed 2 μg.
2. Incubate the reaction system at 50°C for 20 minutes;
3. After the reaction is completed, add an appropriate amount of proteinase K or add 5% volume of 0.5 M EDTA to terminate the reaction.
Precautions
- The optimum reaction temperature of this product is higher than 65°C, and it is still active at 95°C, and its activity at 65°C is 3-4 times that at 37°C.
- All enzymes should be stored in an ice box and stored at -20°C immediately after use.
- For your safety and health, please wear a lab coat and disposable gloves for operation.
