Chinese Science Bulletin

, Volume 43, Issue 10, pp 863–867 | Cite as

Molecular cloning and expression of Pfu DNA polymerase gene and its application in long-distance PCR

  • Li Bing 
  • Gu Cheng 
  • Zhao Jindong 


A 2.3 kb DNA fragment containing Pfu DNA polA gene was amplified by PCR from total DNA ofPyrococcus furiosus and cloned into a pGEM-T vector. The recombinant clone pT-pfu was digested with Nco I and Xho I and the fragment was inserted into an expresion vector pET3d-X. The Pfu polA gene was expressed inEscherichia coli BL21 (DE3). The gene product (Pfu) was purified with heat denaturation, polyethyleneminc (PEI) precipitation and Bio-rex 70 ion-exchange chromatography. The recombinant Pfu was verified by Protein N-terminal sequencing. With the recombinant Pfu. large λ DNA fragments were successfully amplified in long-distance PCR.


Pfu enzyme expression long-distance PCR 


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Copyright information

© Science in China Press 1998

Authors and Affiliations

  • Li Bing 
    • 1
  • Gu Cheng 
    • 1
  • Zhao Jindong 
    • 1
  1. 1.College of life SciencesPeking UniversityBeijingChina

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