Advantages of Thermococcus kodakaraenis (KOD) DNA polymerase for PCR-mass spectrometry based analyses
Advantages of Thermococcus kodakaraenis (KOD) DNA polymerase for PCR-mass spectrometry based analyses
复制标题
DOI：
10.1016/s1044-0305(03)00148-x
复制
发表时间：
2003-06-01
影响因子：
3.2
通讯作者：
Muddiman, DC
中科院分区：
文献类型：
--
作者：
Benson, LM;Null, AP;Muddiman, DC
证明了来自Thermococcus kodakaraensis（KOD）的热稳定DNA聚合酶用于PCR扩增以及随后的质谱检测的优点。用于PCR扩增的常用DNA聚合酶包括来自水生栖热菌（Taq）和激烈火球菌（Pfu）的那些。一个116个碱基对的PCR产物来自一个vWA基因座扩增的Taq，Pfu，或KOD DNA聚合酶和比较琼脂糖凝胶电泳和电喷雾电离傅立叶变换离子回旋共振质谱（ESI-FTICR-MS）。与Pfu或Taq相比，KOD DNA聚合酶的PCR产物形成分别增加了2至3倍，并产生了平端PCR产物，这使得质谱易于解释。此外，我们证明了使用高磁场的优势，以获得相同的116个碱基对（类似于72 kDa）的PCR产物在高m/z的单位分辨率。（J Am Soc Mass Spectrom 2003，14，601 -604）（C）2003年美国质谱学会。
The advantages of the thermostable DNA polymerase from Thermococcus kodakaraensis (KOD) are demonstrated for PCR amplification with subsequent detection by mass spectrometry. Commonly used DNA polymerases for PCR amplification include those from Thermus aquaticus (Taq) and Pyrococcus furiosus (Pfu). A 116 base-pair PCR product derived from a vWA locus was amplified by Taq, Pfu, or KOD DNA polymerase and compared by agarose gel electrophoresis and electrospray ionization Fourier transform ion cyclotron resonance mass spectrometry (ESI-FTICR-MS). KOD DNA polymerase demonstrated a 2- to 3-fold increase in PCR product formation compared to Pfu or Taq, respectively, and generated blunt-ended PCR product which allows facile interpretation of the mass spectrum. Additionally, we demonstrate the advantage of using high magnetic fields to obtain unit resolution of the same 116 base pair (similar to72 kDa) PCR product at high m/z. (J Am Soc Mass Spectrom 2003,14,601-604) (C) 2003 American Society for Mass Spectrometry.