Polymerase enhancing factor (PEF) extracts, PEF protein complexes, isolated PEF proteins, and methods for purifying and identifying them
원문보기
IPC분류정보
국가/구분
United States(US) Patent
등록
국제특허분류(IPC7판)
C12P-019/34
C12P-019/00
출원번호
UP-0631613
(2000-08-04)
등록번호
US-7625725
(2009-12-16)
발명자
/ 주소
Hogrefe, Holly
Hansen, Connie J.
출원인 / 주소
Stratagene California
인용정보
피인용 횟수 :
0인용 특허 :
37
초록▼
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 compo
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 Pyrococcus furiousus PEF (P45), which was used to produce a recombinant PEF.
대표청구항▼
What is claimed is: 1. A method of enhancing a nucleic acid polymerase reaction comprising: (a) forming a nucleic acid polymerase reaction composition comprising: (i) a nucleic acid (ii) at least one nucleic acid polymerase selected from a Pfu DNA polymerase, an exo-Pfu DNA polymerase, a Pwo DNA po
What is claimed is: 1. A method of enhancing a nucleic acid polymerase reaction comprising: (a) forming a nucleic acid polymerase reaction composition comprising: (i) a nucleic acid (ii) at least one nucleic acid polymerase selected from a Pfu DNA polymerase, an exo-Pfu DNA polymerase, a Pwo DNA polymerase, a Vent DNA polymerase, a Deep Vent DNA polymerase, a JDF3 DNA polymerase, and an ES4 DNA polymerase, and (iii) a P45 protein, wherein the P45 protein is in monomeric, dimeric, or multimeric form, and wherein the P45 protein is produced from a cell containing a DNA construct comprising a sequence encoding polymerase enhancing factor protein P45 operably linked to an expression vector, and wherein the P45 protein comprises the sequence of SEQ ID NO:72, and (b) incubating the nucleic acid polymerase reaction composition under conditions allowing a nucleic acid polymerase reaction, wherein the P45 protein increases rate, fidelity and/or yield of the nucleic acid polymerase reaction. 2. A method of enhancing a nucleic acid polymerase reaction as claimed in claim 1, wherein the P45 protein is present in a polymerase enhancing factor complex. 3. A method for controlling the activity of a polymerase in a nucleic acid polymerase reaction, comprising: (a) forming a nucleic acid polymerase reaction composition comprising: (i) a nucleic acid (ii) at least one nucleic acid polymerase selected from a Pfu DNA polymerase, an exo-Pfu DNA polymerase, a Pwo DNA polymerase, a Vent DNA polymerase, a Deep Vent DNA polymerase, a JDF3 DNA polymerase, and an ES4 DNA polymerase, and (iii) a polymerase enhancing factor activity, wherein the polymerase enhancing factor activity comprises the sequence of SEQ ID NO:72, and wherein the the polymerase enhancing activity changes the amount of dUTP present or generated during the reaction, and (b) incubating the nucleic acid polymerase reaction composition under conditions allowing a nucleic acid polymerase reaction, wherein changing the amount of dUTP present or generated during the reaction controls the activity of the polymerase in the polymerization reaction. 4. A method according to claim 3, wherein the polymerase enhancing factor activity comprises a P45 protein, wherein the P45 protein is in monomeric, dimeric, or multimeric form, and wherein the P45 protein is produced from a cell containing a DNA construct comprising a sequence encoding polymerase enhancing factor protein P45 operably linked to an expression vector.
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이 특허에 인용된 특허 (37)
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Holly Hogrefe, Polymerase enhancing factor (PEF) extracts, PEF protein complexes, isolated PEF proteins, and methods for purifying and identifying same.
Holly Hogrefe, Polymerase enhancing factor (PEF) extracts, PEF protein complexes, isolated PEF proteins, and methods for purifying and identifying same.
Comb Donald G. (Beverly MA) Perler Francine (Brookline MA) Kucera Rebecca (Beverly MA) Jack William E. (Rowley MA), Purified thermostable DNA polymerase obtainable from thermococcus litoralis.
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Comb Donald G. (Beverly MA) Perler Francine (Brookline MA) Kucera Rebecca (Beverly MA) Jack William E. (Wenham MA), Recombinant thermostable DNA polymerase from archaebacteria.
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Gelfand David H. (Oakland CA) Myers Thomas W. (Emeryville CA), Reverse transcription with thermostable DNA polymerase-high temperature reverse transcription.
Barnes Wayne M. (223 Renaldo Dr. Chesterfield MO 63017), Thermostable DNA polymerase with enhanced thermostability and enhanced length and efficiency of primer extension.
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