Taq-Polymerase

• UniProt: P19821

Taq polymerase is a thermostable DNA polymerase from the bacterium Thermus aquaticus (Taq ). This bacterium lives in geysers at about 70 ° C. Taq polymerase is extremely heat resistant and is one of the Extremozymen. The reason for this is the presence of a mutated cold shock protein, which differs from similar proteins of mesophilic bacteria through the exchange of two amino acids. In 1969, the Taq polymerase was first isolated by Thomas Brock and Hudson Freeze.

The enzyme is mainly used for DNA amplification using the polymerase chain reaction. DNA polymerases from mesophilic organisms were denatured when heated during the polymerase chain reaction and should be re- added after each cycle. In DNA amplification with Taq polymerase, this is not necessary, as the enzyme even at high temperatures, there is very stable ( the enzyme half-life at 97.5 ° C for about 9 minutes). The DNA amplification with Taq is error -prone, because the enzyme has no proofreading function. In the amplified DNA fragments therefore find mutations that are due to inaccurate copying the Matritzenstranges. If exact sequence DNA amplicons required, hence the use of other polymerases, such as Pfu or Pfx polymerase (which were also originally isolated from thermophilic microorganisms), their use, however, is more costly recommended.

In the amplification of a DNA fragment, the Taq polymerase, a depends additional nucleotide ( dATP ) to the 3 ' end of the synthesized strand. One speaks of a 3' -A overhang (A for adenine), since there is no complementary nucleobase ( thymine ) on the template strand. The 3'-overhang is not corrected by the lack of proofreading function and applies the process of the TA- cloning.

The mass production of the enzyme occurs by transfer of the gene to a strain of the bacterium Escherichia coli. From these cultures, the protein is purified and stored in a buffer containing glycerol at -20 ° C.