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Original ArticleOpen Access
Production and Evaluation of Taq DNA Polymerase
Leelayuwat C 
,
Srisuk T ,
Paechaiyaphum R ,
Limpaiboon T ,
Romphruk A ,
Romphruk A
,
Srisuk T ,
Paechaiyaphum R ,
Limpaiboon T ,
Romphruk A ,
Romphruk A Taq DNA polymerase is an enzyme essential in performing Polymerase Chain Reaction
(PCR) which has recently become a basic technology in research and diagnostic laboratories. In
order to reduce the cost of research work in Thailand, recombinant Taq DNA polymerase was
locally produced from pTaq cloned in E. coli. The enzyme was characterized and evaluated in comparison
with the commercial Taq DNA polymerase produced by Perkin Elmer Cetus, U.S.A. The
yield of enzyme was 6.72 mg/ml and the activity of 9,524 units/mg protien with the total of 448,000
units/litre of the bacterial culture. The preparation was free of DNase based upon its ability to degrade
Lambda DNA evaluated by gel electrophoresis. Although the enzyme produced gave a high DNA
polymerase activity, the preparation was not as pure as the enzyme produced by Perkin Elmer
Cetus. Immunoblot analysis indicated that the enzyme preparation contained the products of
enzyme degradation obtained during preparation and bacterial protein contaminations. In spite
of the existence of bacterial proteins in the preparation, the Taq enzyme produced was proved to
be applicable in performing PCR such as the PCR-SSP (Sequence Specific Primers) typing for
HLA-DR. The cost of enzyme preparation was about 256 times less than that of the commercial
enzyme. Economically, the locally produced Taq DNA polymerase can be used efficiently in the
research laboratories performing PCR based typing of the HLA genes.
(PCR) which has recently become a basic technology in research and diagnostic laboratories. In
order to reduce the cost of research work in Thailand, recombinant Taq DNA polymerase was
locally produced from pTaq cloned in E. coli. The enzyme was characterized and evaluated in comparison
with the commercial Taq DNA polymerase produced by Perkin Elmer Cetus, U.S.A. The
yield of enzyme was 6.72 mg/ml and the activity of 9,524 units/mg protien with the total of 448,000
units/litre of the bacterial culture. The preparation was free of DNase based upon its ability to degrade
Lambda DNA evaluated by gel electrophoresis. Although the enzyme produced gave a high DNA
polymerase activity, the preparation was not as pure as the enzyme produced by Perkin Elmer
Cetus. Immunoblot analysis indicated that the enzyme preparation contained the products of
enzyme degradation obtained during preparation and bacterial protein contaminations. In spite
of the existence of bacterial proteins in the preparation, the Taq enzyme produced was proved to
be applicable in performing PCR such as the PCR-SSP (Sequence Specific Primers) typing for
HLA-DR. The cost of enzyme preparation was about 256 times less than that of the commercial
enzyme. Economically, the locally produced Taq DNA polymerase can be used efficiently in the
research laboratories performing PCR based typing of the HLA genes.
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