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Description
Thermo Scientific RevertAid Reverse Transcriptase (RT) is a recombinant M-MuLV RT. It differs from the M-MuLV RT by its structure and catalytic properties. The enzyme possesses an RNA-dependent and DNA-dependent polymerase activity and a RNase H activity specific to RNA in RNA-DNA hybrids that is significantly lower than that of Avian Myeloblastosis Virus (AMV) reverse transcriptase.
Features of RevertAid Reverse Transcriptase include:
• Efficient synthesis of full-length first strand cDNA up to 13 kb
• Optimum activity at 42°C
• Active up to 50°C
• Incorporates modified nucleotides (e.g., Cy3-, Cy5-, rhodamine-, aminoallyl-, and fluorescein-labeled nucleotides)
Applications
• First strand cDNA synthesis for RT-PCR and real-time RT-PCR
• Synthesis of cDNA for cloning and expression
• Generation of labeled cDNA probes for microarrays
• DNA labeling
• Analysis of RNA by primer extension
Source
RevertAid Reverse Transcriptase is sourced from E.coli cells with a cloned fragment of the pol gene encoding Moloney Murine Leukemia Virus reverse transcriptase.
Specifications
Specifications
| Concentration | 200 U/μL |
| Content And Storage | • RevertAid Reverse Transcriptase (200 U/μL) |
| Format | Stand-alone Enzyme |
| Reaction Speed | 60 min. |
| Technique | Reverse Transcription |
| Optimal Reaction Temperature | 42°C |
| Quantity | 5 x 10,000 Units |
| Reverse Transcriptase | RevertAid |
| Ribonuclease H Activity | Yes |
| Shipping Condition | Dry Ice |
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Frequently Asked Questions (FAQs)
It is generally beneficial to minimize RNase H activity when aiming to produce long transcripts for cDNA cloning. RNase H degrades RNA from RNA-DNA duplexes, which can result in truncated cDNA during reverse transcription of long mRNA. We also recommended using RNase H-minus RTs for template-independent addition of C nucleotides.
All Thermo Scientific reverse transcriptases possess intrinsic TdT activity although at varying degrees depending upon the reaction conditions. For addition of template-independent C nucleotides (as for SMART and RACE experiments), this specific TdT activity can be induced by Mn2+. We would recommend Maxima H- or RevertAid H- minus RTs for this purpose.
Trace amounts of reagents used in RNA purification protocols may remain in solution and inhibit first-strand synthesis, e.g., SDS, EDTA, guanidine salts, phosphate, pyrophosphate, polyamines, spermidine. To remove trace contaminants, we recommend re-precipitating the RNA with ethanol and washing the pellet with 75% ethanol, or re-purifying the RNA.
RNA purity and integrity are essential for synthesis and quantification of cDNA. Always assess the integrity of RNA prior to cDNA synthesis. Use freshly prepared RNA. Multiple freeze/thaw cycles of the RNA sample and synthesized cDNA is not recommended. Avoid RNase contamination and discard low quality RNA.
It is generally beneficial to minimize RNase H activity when aiming to produce long transcripts for cDNA cloning. RNase H degrades RNA from RNA-DNA duplexes, which can result in truncated cDNA during reverse transcription of long mRNA. We also recommended using RNase H Minus RTs for template-independent addition of C nucleotides. In contrast, reverse transcriptases with intrinsic RNase H activity are often favored in 1-step RT-qPCR applications.
For Research Use Only. Not for use in diagnostic procedures.
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