Pfu replication accessory factors and methods of use

Abstract

This invention provides isolated polynucleotides that encode replication accessory factors. The invention also provides novel DNA replication accessory factors, which have been isolated and purified from the hyperthermophilic archaea Pyrococcus furiosus . The invention also provides various methods of enhancing a nucleic acid polymerase reaction comprising the addition of the replication accessory factors to the reaction. This invention further provides methods of synthesizing, amplifying, and mutagenizing nucleic acids of interest employing the replication accessory factors. This invention also provides kits comprising, at least one of the replication accessory factors. This invention also provides kits useful for various methods that comprise at least one replication accessory factor.

1 - 3 . (canceled) 
     
     
         4 . An isolated polynucleotide encoding a helicase, wherein the polynucleotide comprises a nucleic acid sequence that encodes an amino acid sequence at least 70% identical to SEQ ID NO:74. 
     
     
         5 . The polynucleotide of  claim 4 , wherein the polynucleotide is a cDNA. 
     
     
         6 . The polynucleotide of  claim 4 , wherein the polynucleotide is an mRNA. 
     
     
         7 - 14 . (canceled) 
     
     
         15 . A vector comprising the polynucleotide of  claim 4 . 
     
     
         16 . The vector of  claim 15 , wherein the vector is a plasmid. 
     
     
         17 . The vector of  claim 15 , wherein the vector is a bacteriophage. 
     
     
         18 . The vector of  claim 15 , wherein the vector is a retrovirus. 
     
     
         19 . The vector of  claim 15 , wherein the vector is an adenovirus. 
     
     
         20 . A host cell comprising the vector of  claim 15 . 
     
     
         21 . The host cell of  claim 20 , wherein the host cell is a prokaryotic cell. 
     
     
         22 . The host cell of  claim 20 , wherein the host cell is a eukaryotic cell. 
     
     
         23 - 26 . (canceled) 
     
     
         27 . A method for producing a polypeptide, comprising: expressing the polynucleotide of the vector of  claim 15  in a host cell; and purifying the expressed product. 
     
     
         28 . The method of  claim 27 , wherein the host cell is a prokaryotic cell. 
     
     
         29 . The method of  claim 27 , wherein the host cell is a eukaryotic cell. 
     
     
         30 - 74 . (canceled) 
     
     
         75 . The polynucleotide of  claim 4 , wherein the amino acid sequence is at least 80% identical to SEQ ID NO:74. 
     
     
         76 . The polynucleotide of  claim 4 , wherein the amino acid sequence is at least 90% identical to SEQ ID NO:74. 
     
     
         77 . The polynucleotide of  claim 4 , wherein the amino acid sequence is at least 95% identical to SEQ ID NO:74. 
     
     
         78 . The polynucleotide of  claim 4 , wherein the nucleic acid sequence comprises SEQ ID NO:67.