Methods (molecular, IHC, frozen)
Molecular
PCR
Taq polymerase
Topic Completed: 1 July 2010
Minor changes: 15 February 2019
Copyright: 2008-2018, PathologyOutlines.com, Inc.
PubMed Search: Taq polymerase[TI]
Table of Contents
Definition / generalCite this page: Shackelford R. Taq polymerase. PathologyOutlines.com website. https://www.pathologyoutlines.com/topic/molecularpathtaqpolymerase.html. Accessed January 18th, 2021.
Definition / general
- Initially the Klenow fragment from the E. coli DNA polymerase I was used in PCR reactions
- While the Klenow fragment worked in PCR, it rapidly denatured at the 90 °C or higher temperatures required for denaturation, requiring the addition of new Klenow polymerase after every denaturation step
- The need to open the reaction chamber with every PCR thermocycle made PCR cumbersome and greatly increased the chances for contamination with unwanted DNA
- The solution to this problem came from the discovery and cloning of a thermostable DNA polymerase from the thermophilic bacterium Thermus aquaticus
- The polymerase, known as the Taq polymerase, has many attributes that are well suited for PCR:
- Enzyme works best at 75 - 80 °C, allowing the elongation step to occur at temperatures which make non-Watson-Crick base paring a rare event
- Its half life at 95 °C is over 40 minutes
- At 72 °C it can add 1,000 nucleoside triphosphates to a growing DNA strand
- Most PCR reactions done with Taq can be completely closed through all the thermocycles
- Taq lacks 3' to 5' exonuclease proofreading activity, allowing a roughly 1 per 9,000 error rate in nucleoside triphosphate incorporation
- Additionally, Taq often adds an adenine to the end of its polymerase reaction (sometimes used to facilitate TA cloning)
- Taq was named "The Molecule of the Year" by Science in 1989
