PCR and PCR type testing is a rapidly expanding market. No longer just an experiment that is performed in research laboratories, you can find PCR apparatus in your physician's office, in use at breweries and dairy farms and even kits for home-based testing to rule out allergens that could provoke an anaphylactic shock reaction.
Visitors shopping for laboratory supplies on our website frequently ask us questions about the different Taq and Master Mix formulations we offer, and so we thought it would be helpful to ask an expert to assist us with an article about one variation of Taq and how this manufacturer certifies their products to be reliable for PCR.
Thank you Susan Johnson from Accuris Instruments for making time to share this with us.
Taq Polymerase was first isolated in 1976 and found its use in the mid 80’s after the development of PCR by Kerry Mullis. Mullis won the Nobel prize in chemistry for his work with PCR in 1993, but Taq Polymerase received Science Magazine’s “Molecule of the Year” honors four years earlier, in 1989.
While Taq was (and is) a great tool in the molecular biology laboratory, it is not without its problems.
One of these problems is that the enzyme exhibits some activity at room temperature and even when on ice. This can result in non-specific amplification and the formation of primer-dimers during reaction set up.
To remedy this issue, researchers chemically modified specific amino acid residues to the Taq Polymerase. This modification blocks the enzyme activity until the reaction mixture is heated to 95ºC for 10-15 minutes. This was a huge innovation in increasing enzyme specificity.
But, while Taq Polymerase is a thermostable enzyme, its half-life is significantly decreased when the enzyme is exposed to temperatures above 90ºC.
This can be resolved by adding an excess on enzyme, but it does not lessen the wait time of 10-15 minutes for a reaction that might only take an hour or so to complete.
More recently, a different way of tying up the active sites of the enzyme has been developed – antibody mediated hot start.
In this process, a monoclonal antibody binds to the Taq Polymerase, preventing enzyme activity and DNA synthesis.
It takes less than a minute at 95º to denature the antibody and restore the activity of Taq Polymerase, solving both the half-life and time sink issues.
Accuris Hot Start Taq is an antibody mediated polymerase.
A little more information about antibody mediated polymerases and Accuris Hot Start Taq:
- Antibody mediated Taq utilizes a monoclonal antibody purified from mice. (Note that some labs that may be using PCR for human or mammalian diagnostic testing might not want the presence of any antibodies in their samples and may not want to use antibody mediated Taq.)
- The process of purifying the mouse monoclonal antibody must be very strict as there is a slight possibility of contamination
- Quality control is important with any type of Taq used, and Accuris offers the highest level of QC available
- Each production batch of Accuris Taq is produced and purified under strict guidelines. Prior to final aliquoting, Accuris Taq is tested for both mammalian and bacterial DNA contamination using qPCR and SOX9, B2M and Gamma Actin primers
- Accuris Hot Start Taq is available either as a stand alone product supplied with an optimized buffer; as a 2X Master Mix, or 2X Master Mix with Red Loading Dye already added.
When lab supply shopping for PCR plates, PCR strip tubes and PCR Master Mixes, Stellar Scientific has a robust catalog and expert product knowledge to help you make the right choice.