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Thermostable Reverse Transcriptase

Ordering

Cat. No.
Size
BIO-65111
10,000 Units
BIO-65112
4 x 10,000 Units

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Description

Thermostable Reverse Transcriptase is a robust thermostable MMLV-RT, designed to reduce RNase H activity and increase thermal stability. The enzyme offers higher cDNA yields and improved efficiency on GC-rich target RNAs and RNA with high secondary structure, such as viral RNA sequences.

Product Highlights

  • Stable - Reverse transcriptase activity up to 60°C
  • Robust - Ideal for RNA with high secondary structure such as viral genomes
  • Sensitive - Detection of low copy number RNA targets
  • Fast - Ideal for developing fast, highly reproducible RT-qPCR assays

Product Description

Traditional Moloney Murine Leukemia Virus Reverse Transcriptase (MMLV-RT) is not thermostable and can only maintain its enzymatic activity at relatively low temperatures (up to 50°C). However for cDNA synthesis, a higher reaction temperature is desirable as it reduces RNA secondary structures which can inhibit reverse transcription and it minimizes nonspecific primer binding.

Meridian Thermostable Reverse Transcriptase has higher thermal stability and reduced RNase H activity. The enzyme can be used to synthesize first-strand cDNA at temperatures up to 60°C which improves the cDNA yield from difficult RNA targets that require higher temperature to denature strong secondary structures.

Main

Real-Time PCR Selection Chart


Specification

Components

Reagent

10,000 units

4 x 10,000 units

Thermostable Reverse Transcriptase

50 µL

4 x 50 µL

5x Reaction Buffer

1 mL

4 x 1 mL

Storage & Stability

Thermostable Reverse Transcriptase can be stored for up to 12 months at -20°C. Repeated freeze/thaw cycles should be avoided.

When stored under the recommended conditions and handled correctly, full activity of the kit is retained until the expiry date printed on the outer box label.

Shipping conditions

On dry or blue ice



Resources

Documents

Thermostable Reverse Transcriptase - Fact Sheet
Thermostable Reverse Transcriptase - Product Manual
Thermostable Reverse Transcriptase - MSDS

FAQs

I want to generate cDNA from RNA with very high secondary structure, can I use Thermostable Reverse Transcriptase?

Yes, for RNA with very high secondary structure we recommend using Thermostable Reverse Transcriptase as it will work at temperatures up to 60°C, which improves the cDNA yield from difficult RNA targets that require higher temperature to denature strong RNA secondary structures.

Is Thermostable Reverse Transcriptase RNase H minus?

Thermostable Reverse Transcriptase does contain the domain for RNase H, but it has been inactivated, so that there is no detectable RNase H activity.

Should I treat the cDNA with RNase H prior to further downstream processing?

If you are going to do RT-PCR or RT-qPCR, this is unnecessary, RNA:DNA duplex after cDNA synthesis are melted during the 95°C denaturing step at the start of the PCR reaction. For cloning of larger fragments, RNase H treatment can be beneficial.

How is Thermostable Reverse Transcriptase inactivated?

The reverse transcriptase can be inactivated by adding a chelating agent such as EDTA or heating to 70°C or higher for 10 min.

Can Thermostable Reverse Transcriptase be used for long transcripts?

Thermostable Reverse Transcriptase can be used for first-strand cDNA synthesis at temperatures up to 60°C, providing increased specificity, higher yields and can generate cDNA up to 12 kb.

Does Thermostable Reverse Transcriptase need a primer?

Yes, to initiate reverse transcription, the MMLV-RT reverse transcriptase require a primer to bind to its complementary sequences on the RNA template and serve as a starting point for synthesis of a new strand. For general cDNA synthesis, random hexamer primers and oligo(dT) primers can be used either separately or mixed. For RT-PCR and RT-qPCR gene-specific primers should be used.

What is the error rate of Thermostable Reverse Transcriptase?

Thermostable Reverse Transcriptase -based reverse transcriptase has an error rate in the range of about 4.8 × 10−5.

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