Purified thermostable enzyme

• US 4,889,818 A
• Filed: 06/17/1987
• Issued: 12/26/1989
• Est. Priority Date: 08/22/1986
• Status: Expired due to Term

• Current Assignee: Roche Molecular Systems (Roche Holding AG)
• Original Assignee: Cetus Corporation (Bayer AG)
• Inventors: Gelfand, David H., Saiki, Randall K., Stoffel, Susanne, Lawyer, Frances C.
• Primary Examiner(s): Wiseman, Thomas G.
• Assistant Examiner(s): Carson, Patricia
• Application Number:
  US07/063,509
• Time in Patent Office: 923 Days
• Field of Search: 435/183, 435/194, 935/14
• US Class Current: 435/194
• CPC Class Codes: C12N 9/1252 DNA-directed DNA polymerase (2.7.7.7), i.e. DNA replicase
• CPC Class Codes: C12N 9/1276 RNA-directed DNA polymerase (2.7.7.49), i.e. reverse transcriptase or telomerase
• CPC Class Codes: C12P 19/34 Polynucleotides, e.g. nucleic acids, oligoribonucleotides

Abstract

A purified thermostable enzyme is obtained that has unique characteristics. Preferably the enzyme is isolated from the Thermus aquaticus species and has a molecular weight of about 86,000-90,000 daltons. The thermostable enzyme may be native or recombinant and may be used in a temperature-cycling chain reaction wherein at least one nucleic acid sequence is amplified in quantity from an existing sequence with the aid of selected primers and nucleotide triphosphates. The enzyme is preferably stored in a buffer of non-ionic detergents that lends stability to the enzyme.

Claims

1. Purified thermostable Thermus aquaticus DNA polymerase that migrates on a denaturing polyacrylamide gel faster than phosphorylase B and more slowly than does bovine serum albumin and has an estimated molecular weight of 86,000-90,000 daltons when compared with a phosphorylase B standard assigned a molecular weight of 92,500 daltons

2. The polymerase of claim 1 that is isolated from Thermus acquaticus.

3. The polymerase of claim 1 that is isolated from a recombinant organism transformed with a vector that codes for the expression of Thermis aquaticus DNA polymerase.