myPOLS Biotec is coming from a scientific tradition: Our expertise lies especially in the development of DNA polymerase assays and in the engineering of DNA polymerases with new properties and functions. The Volcano product family is based on a polymerase engineered by our scientists.
Special products for special requirements
All products of the Volcano family are applicable for reverse transcription reactions and have a thermostable nature. We recommend to design primers to have higher melting/annealing temperatures (above 65°C). Additionally, for assay setup it is advised to perform RT-PCR test reactions with temperature gradients (i.e., for the annealing and extension steps) to identify the most proficient PCR protocol for your needs using our Volcano DNA polymerase products.
Depending on the employed assay, the reverse transcription steps may be omitted (0-step RT-PCR).
Some practical advice establishing RT-PCR assays with RevTaq RT-PCR DNA polymerase or Volcano3G® RT-PCR Master Mixes:
1. Make sure that your primers have melting points higher than 65°C.
2. Optimize PCR-steps: Prepare one big PCR reaction with cDNA and aliquot it into the plate in multiple wells. Apply a two-step PCR protocol (for example 95°C 10 sec. and a temperature gradient between 57-67°C (1min, 40 cycles).
> From this experiment you can select the best temperature/well which shows the earliest Ct-value.
3. Optimize cDNA generation/reverse transcription steps: Then, take an RNA sample with a relatively high amount of target RNA (~107-105 copies/rxn) and run an RT-PCR with a protocol such as this: 10 cycles of 95°C 3sec and a temperature gradient between 60-75 °C for 1 min) and followed by PCR cycles with the best PCR temperature as selected in step 2.
> From this experiment you can select the best temperature/well with the earliest Ct-value.
These two steps above combined should already result in a very decent RT-PCR protocol. Finetuning is possible by shortening the extension-times during PCR phase or further optimizations of the denaturation temperatures (via temperature gradient RT-PCRs e.g. between 85-100°C).
Further development to a new generation of reverse transcriptase - let's do it together
You want to speed up your existing RT-PCR assays or looking for optimized components to develop a new test? Want to have more sensitivity? Let´s do this together. The in-house development facilitates further development. We know our enzymes. This enables us to respond to customer requirements and to adapt the products ideally to specific customer needs and targets.
For example: Some products are already successfully used in IVD products for the pathogenic detection of RNA viruses, like SARS-CoV-2.
These Volcano components are interesting for you if you want to establish new tests or optimize existing tests with new procedures - for research only use and diagnostic purpose.
RevTaq RT-PCR DNA polymerase: the pure, reverse transcription active enzyme
RevTaq RT-PCR DNA polymerase is an engineered, extremely thermostable reverse transcriptase and combined DNA polymerase, obtained through directed, artificial evolution. (to the shop)
Volcano3G® RT-PCR Master Mixes: different variants for different needs
Volcano3G® RT-PCR Master Mix products contain all components required for general RT-qPCR. The products are offered for research use only and for in-vitro diagnostic purpose.
Direct SARS-CoV-2 RT-PCR detection kit w/o RNA extraction
The kit contains Volcano3G® RT-PCR Probe 2x Master Mix and a duplex primer/probe mix for both SARS-CoV-2 (N1 gene specific) and RNaseP control. Please also refer the procedure as detailed in our recent reference: "Detection of SARS-CoV-2 from raw patient samples by coupled high temperature reverse transcription and amplification" (to the shop)
Find all Volcano products at a glance (to the shop)
Interested? You can contact us for a special quote or more details using our contact form. Products for research only use can be purchased through the shop.