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I left the RapiDxFire Thermostable Reverse Transcriptase (RT) out on the bench overnight. Is it still good?

307

Yes, the RapiDxFire RT is stable at room temperature (20 °C) for at least three months. However, we have not tested the stability of the combined reagents (buffer, template, primers) over time at room temperature.

What is the longest cDNA product that RapiDxFire Thermostable RT can synthesise?

307

RapiDxFire Thermostable RT has successfully synthesised products up to 1kb in 5 minutes.

How should I extract or purify my RNA samples if I wish to reverse transcribe them using EpiScript RNase H- Reverse Transcriptase?

221, RDXERT300

MasterPure™, sbeadex™ and QuickExtract™ isolation chemistries are all suitable for use with EpiScript RNase H- Reverse Transcriptase.

How should I extract or purify my RNA samples if I wish to reverse transcribe them using EpiScript RT PLUS?

AS3107, MBTOOL-007, PCRAMP-104

MasterPure™, sbeadex™ and QuickExtract™ isolation chemistries are all suitable for use with EpiScript RT PLUS.

Can I shorten the activation time for the heat-activated thermal stable reverse transcriptase?

PCRAMP-021

Shortening the activation time for the heat-activated reverse transcriptase is not recommended. We recommend a three-minute activation time to fully activate the reverse transcriptase. However, assay optimisation may enable shortening the activation time to a minimum of one minute depending on other factors such as other thermocycling parameters, sample type, and sensitivity requirements.

What is the highest temperature the cDNA synthesis step can be performed?

PCRAMP-021

The highest temperature for cDNA synthesis is partly dependent on TM of the primers used. Other factors include the length of the desired amplicon and if there is potential RNA secondary structure.

What is the recommended concentration for EpiScript RNase H- Reverse Transcriptase in RT reactions?

221, RDXERT300

For 1-step RT-qPCR, we have found 20-50 units per reaction to work best. In 2-step RT-qPCR, we have found 5-20 units per reaction to work best.

What conditions do you recommend for set up of RT reactions using EpiScript RNase H- Reverse Transcriptase?

221, RDXERT300

For best results we recommend preparing reactions on ice for both 1-step and 2-step RT-qPCR experiments.

Can I use random hexamer or nonamers with EpiScript RNase H- Reverse Transcriptase?

221, RDXERT300

Random primers can be used with EpiScript RNase H- Reverse Transcriptase. If using random primers, we suggest that the RT step is performed at 42 °C or lower as many random primers will not anneal at higher temperatures.

Is EpiScript RNase H- Reverse Transcriptase suitable for RNA-seq library preparation?

221, RDXERT300

Yes, EpiScript RNase H- Reverse Transcriptase is compatible with RNA-Seq.