Optimization of PCR with deep vent DNA polymerase for amplification of NS4 gene - hepatitis C virus
- Author:
Giang Hoai Nguyen
- Publication Type:Journal Article
- Keywords:
Hepatitis C
- MeSH:
Hepatitis C;
Deep Vent DNA polymerase
- From:Journal of Medical Research
2002;17(12):9-12
- CountryViet Nam
- Language:Vietnamese
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Abstract:
Deep vent DNA polymerase is purified from a strain of E. Coli that carries the Deep Vent DNA polymerase gene from pyrococcus species GB-D. The native organism was isolated from a submarine thermal vent at 2010 meters and is able to grow at temperature as high as 104oC. Deep Vent DNA polymerase is a high-fidelity thermophilic DNA polymerase. The Deep Vent DNA polymerase is derived from an integral 3'-5' proofreading exonuclease activity. Deep Vent DNA polymerase is ven more stable than Vent DNA polymerase at temperature of 95 to 100oC. Because of these advantages, we want to apply Deep Vent DNA polymerase for amplification of NS4 gene - hepatitis C virus by PCR. Our study showed that PCR conditions for Deep Vent DNA polymerase are rather different than for Taq DNA polymerase. The amplification of NS4 gene - hepatitis C virus with Deep Vent DNA polymerase is optimal with the following parameters: (1) A final concentration of MgSO4 in PCR mixture of 4 mM. (2) An annealing temperature of 57oC. (3) A final concentration of dNTPs of 200 mM.
