Patent ID: 6395526
Filing Date: 2002-05-28
Classification: C12N

Abstract:
An isolated and purified DNA polymerase, characterized in that said DNA polymerase possesses the following properties:1) exhibiting higher polymerase activity when assayed by using as a substrate a complex resulting from primer annealing to a single stranded template DNA, compared to using an activated DNA as a substrate; 2) possessing a 3â€²â†’5â€² exonuclease activity; 3) amplifying a DNA fragment of about 20 kbp, when polymerase chain reaction (PCR) is carried out using &lgr;-DNA as a template under the following conditions: PCR conditions: (a) a composition of reaction mixture: containing 10 mM Tris-HCl (pH 9.2), 3.5 mM MgCl2, 75 mM KCl, 400 &mgr;M each of dATP, dCTP, dGTP and dTTP, 0.01% bovine serum albumin, 0.1% Triton X-100, 5.0 ng/50 &mgr;l &lgr;-DNA, 10 pmole/50 &mgr;l primer &lgr;1 (SEQ ID NO:8), primer &lgr;11 (SEQ ID NO:9), and 3.7 units/50 &mgr;l DNA polymerase; (b) reaction conditions: carrying out a 30-cycle PCR, wherein one cycle is defined as at 98Â° C. for 10 seconds and at 68Â° C. for 10 minutes; wherein said DNA polymerase comprises a first polypeptide and a second polypeptide, which are non-covalently bonded to form a complex, wherein said first polypeptide comprises: (I) the amino acid sequence encoded by a DNA of SEQ ID NO:1, or (II) an amino acid sequence encoded by a DNA, the complement thereof hybridizing to the DNA of SEQ ID NO:1 under stringent conditions; and wherein said second polypeptide comprises: (I) the amino acid sequence encoded by a DNA of SEQ ID NO:3, or (II) an amino acid sequence encoded by a DNA, the complement thereof hybridizing to the DNA of SEQ ID NO:3 under stringent conditions.