Reverse Transcriptase
We offer a selection of high-quality reverse transcriptase enzymes help you develop highly sensitive assays.
Ideal for High-Throughput
Reverse transcription is the process of transcribing RNA molecules into complementary DNA (cDNA). Moloney Murine Leukemia Virus reverse transcriptase (MMLV-RT) is the most commonly used reverse transcriptase in molecular biology workflows. It is highly sensitive; however, novel engineering has enhanced efficiency, improved thermostability, and increase cDNA yields.
- Highly sensitive, even low concentrations of template RNA
- Thermostability of up to 60°C, from RNA with high secondary structure
- Sensitive detection of low copy number RNA targets
- High-concentration, glycerol-free options for lyophilization
There are several key important features when determining the best reverse transcriptase for your application. The first is RNase H activity, if the RNase H activity of the reverse transcriptase is too high, it can lead to excessive degradation of the RNA template, resulting in incomplete or inefficient cDNA synthesis. Reverse transcriptase with low RNase H activity minimize the nonspecific degradation of RNA molecules in the reaction, improving the specificity and fidelity of cDNA synthesis. The second is processivity, the number of nucleotides incorporated in a single binding event, most engineered reverse transcriptases have improved processivity, (up to 65x greater than wild-type MMLV reverse transcriptase). The third is thermostability, high temperatures help denature RNA that has strong secondary structures and/or high GC content enabling full-length cDNA synthesis and high yields. Normally MMLV-RT performs optimally at 37°C whereas engineered thermostable MMLV-RTs can withstand temperatures up to 55°C without negatively impacting reverse transcription efficiency. Meridian offers several reverse transcriptases, each containing unique product features such as high thermostability or lyophilization compatibility for creating room-temperature stable mixes.
- 55C MMLV-RT
- Lyo-Compatible MMLV-RT
- RNase-Tolerant MMLV-RT
- MMLV-RT
- Glycerol-Free HS Tth DNA Polymerase (HC)
Catalogs & Brochures
- Reagent Solutions for Molecular Diagnostics
- 55C MMLV-RT
- Accelerating Efficiency: A Comprehensive Guide to Essential Reagents for High-Throughput Molecular Diagnostic Assays (MDx)
FAQs
I want to generate cDNA from viral RNA with very high secondary structure, can I use MMLV-RT?
For RNA with very high secondary structure, we recommend using 55C MMLV-RT (MDX117), this reverse transcriptase can be used at temperatures up to 60°C which improves the cDNA yield from difficult RNA targets that require higher temperature to denature strong secondary structures.
Can I improve the yield when using MMLV-RT?
Avoiding ribonuclease contamination is of primary concern since high quality template RNA is required. You can add RNase Inhibitor (MDX056), alternatively we also have RNase-Tolerant MMLV-RT (MDX043) which includes an RNase inhibitor.
What buffer do you recommend using with MMLV-RT?
If you are going to make cDNA, you will require a buffer that contains 0.2 mM each dATP, dCTP, dGTP & dTTP, 10 mM DTT (dithiothreitol), 25 mM KCl, 3.5 mM MgCl2 and 50 mM Tris–HCl (7.5). However, if you want to do RT-qPCR, we recommend 1-Step qPCR Buffer, 4x (MDX034) or Lyo-Ready 1-Step RT-qPCR Buffer (MDX052).
How is MMLV-RT 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.
Does MMLV-RT need a primer?
Yes, to initiate reverse transcription, the MMVL-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 MMLV-RT?
Unlike eukaryotic replicative polymerases, reverse transcriptase lack exonuclease activity and so is error prone. MMLV-based reverse transcriptase has been reported to have an error rate in the range of 1.1 × 10−4 to 4.8 × 10−4.
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