Polymerase Enhancing Factor (PEF) Extracts PEF Protein Complexes Isolated PEF Proteins and Methods for Purifying and Identifying

• US 20120003713A1
• Filed: 03/18/2011
• Published: 01/05/2012
• Est. Priority Date: 03/21/1997
• Status: Abandoned Application

• Current Assignee: Agilent Technologies, Inc.
• Original Assignee: Agilent Technologies, Inc.
• Inventors: Hogrefe, Holly, Borns, Michael, Muhich, Michael, Hansen, Connie
• Application Number:
  US13/051,317
• Publication Number:
  US 20120003713A1
• Time in Patent Office: Days
• US Class Current: 435/188
• CPC Class Codes: C12N 9/1252 DNA-directed DNA polymerase (2.7.7.7), i.e. DNA replicase
• CPC Class Codes: C12N 9/16 acting on ester bonds (3.1)
• CPC Class Codes: C12N 9/78 acting on carbon to nitrogen bonds other than peptide bonds (3.5)
• CPC Class Codes: C12P 19/34 Polynucleotides, e.g. nucleic acids, oligoribonucleotides
• CPC Class Codes: C12Q 1/6846 Common amplification features
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2521/101 DNA polymerase
• CPC Class Codes: C12Q 2521/319 Exonuclease
• CPC Class Codes: C12Q 2527/125 Specific component of sample, medium or buffer

Abstract

The invention provides novel extracts, proteins, and complexes that improve the polymerization activity of nucleic acid polymerases. Included within the aspects of the invention are methods for identifying compositions with a polymerase enhancing activity, methods for purifying and using these compositions, and specific extracts, proteins, and complexes that function to enhance polymerase activity. As an example, specifically described is nucleotide and amino acid sequence information for a Pyrococcus furiosus PEF (P45), which was used to produce a recombinant PEF.

Claims

1. A non-naturally occurring composition of matter comprising at least one component possessing nucleic acid polymerase enhancing activity selected from the group consisting of:

an isolated or purified naturally occurring polymerase enhancing protein obtained from a bacterial, eukaryotic, or Archaeal source;

a wholly or partially synthetic protein having the same amino acid sequence as said naturally occurring protein or an analog thereof possessing polymerase enhancing activity;

polymerase-enhancing mixtures of one or more of said naturally occurring or wholly or partially synthetic proteins;

a polymerase-enhancing protein complex of one or more of said naturally occurring or wholly or partially synthetic proteins; and

a polymerase enhancing partially purified cell extract containing one or more of said naturally occurring proteins.

2-95. -95. (canceled)