Oxidation-resistant recombinant RNase inhibitor
Recombinant RNase Inhibitor ver.2.0 is an oxidation-resistant RNase inhibitor that is highly stable in enzymatic reactions requiring low DTT concentrations, such as in vitro transcription, in vitro translation, or RT-PCR. Derived from porcine liver, Recombinant RNase Inhibitor ver.2.0 displays enhanced oxidation resistance due to a mutation in a cysteine residue that is very sensitive to oxidation, which prevents degradation and inactivation of the enzyme under oxidizing reaction conditions.
Recombinant RNase Inhibitor ver.2.0 is an oxidation-resistant RNase inhibitor that is highly stable in enzymatic reactions requiring low DTT concentrations, such as in vitro transcription, in vitro translation, or RT-PCR. Derived from porcine liver, Recombinant RNase Inhibitor ver.2.0 displays greatly enhanced oxidation resistance due to a mutation in a cysteine residue that is very sensitive to oxidation, which prevents degradation and inactivation of the enzyme under oxidizing reaction conditions.
The enzyme forms a 1:1 complex specifically with RNase A to inhibit RNase activity; however, it is not effective against RNase H activity. The inhibitory reaction is reversible, and the inhibitor can be irreversibly inactivated to restore ribonuclease activity by dissociating the complex with urea or a sulfhydryl reagent. Moreover, unlike other non-protein competitive inhibitors (e.g., nucleotides and inorganic phosphates), it can easily be removed from the reaction system by phenol extraction.
Overview
- Efficient RNAse A inhibitor
- Highly stable against oxidation
- Ideal for applications requiring low DTT concentration, such as in vitro transcription, in vitro translation, or RT-PCR
- Suitable for long-term storage of samples
Recombinant RNase Inhibitor ver.2.0 is highly resistant to oxidation
Figure 1. Comparison of oxidation resistance among different commercially-available recombinant RNase inhibitors. 16S/23S rRNA was mixed with DTT-free buffer, RNase A, increasing amounts of H2O2, and either Recombinant RNase Inhibitor ver.2.0 (Cat. # 2135; purple line), Recombinant RNase Inhibitor (Cat. # 2313; blue line), or a commercially-available, oxidation-resistant murine RNase inhibitor (Company N; green line). After incubation at 37°C for 10 min followed by incubation at 25°C for 10 min, each reaction was centrifuged at 13,000g for 15 min at 4°C. Nucleic acids released into the supernatant were measured as an indicator of RNase A-mediated rRNA degradation using a NanoDrop spectrophotometer. At concentrations above 0.1% H2O2, the use of Recombinant RNase Inhibitor ver.2.0 resulted in less rRNA degradation compared to other RNase Inhibitor products. n=2, where each experiment was performed in triplicate.
More Information
Applications
- In vitro transcription/translation (1 U/µl reaction)*
- In vitro transcription/translation with cell-free extract (20 U/µl reaction)*
- RT-PCR (0.5 U/µl reaction)*
- cDNA synthesis (0.5 U/µl reaction)*
- Polysome isolation (1 U/µl reaction)*
*The parentheses show the recommended concentration for each application. For a unit definition, please refer to Certificates of Analysis under the Documents tab.
Source
Escherichia coli carrying a plasmid containing a mutated gene for ribonuclease inhibitor from porcine liver.
Properties
- Molecular mass: approx. 52 kDa
- Optimal pH: maximum inhibition of RNase A occurs at pH 7–8, though inhibition is observed over a broad pH range
Additional product information
Please see the product's Certificate of Analysis for information about storage conditions, product components, and technical specifications. Please see the Kit Components List to determine kit components. Certificates of Analysis and Kit Components Lists are located under the Documents tab.
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