Solubilized phospholipids for stabilizing nucleic acid polymerases

• US 8,715,987 B2
• Filed: 04/19/2012
• Issued: 05/06/2014
• Est. Priority Date: 05/02/2011
• Status: Active Grant

• Current Assignee: New England Biolabs Incorporated
• Original Assignee: New England Biolabs Incorporated
• Inventors: Johnson, Donald, Evans, Thomas C. Jr.
• Primary Examiner(s): NOAKES, SUZANNE MARIE
• Application Number:
  US13/450,549
• Publication Number:
  US 20120282669A1
• Time in Patent Office: 747 Days
• Field of Search: None
• US Class Current: 435/188
• CPC Class Codes: C12N 9/1241 Nucleotidyltransferases (2.7.7)
• CPC Class Codes: C12N 9/96 Stabilising an enzyme by forming an adduct or a composition; Forming enzyme conjugates
• CPC Class Codes: C12Q 1/686 Polymerase chain reaction [PCR]
• CPC Class Codes: C12Q 2521/101 DNA polymerase
• CPC Class Codes: C12Q 2527/127 the enzyme inhibitor or activator used

Abstract

Compositions and methods are provided that relate to solubilized phospholipids and their use in stabilizing nucleic acid polymerases. For example, a phospholipid with a tail containing at least 8 carbons can be solubilized in the presence of an amphipathic molecule.

Claims

1. A composition comprising:

a thermostable polymerase in a buffer comprising a solubilized long-chain phospholipid, wherein the phospholipid has at least one hydrophobic tail containing 8 to 30 carbons.

2. A composition according to claim 1, further comprising an amphipathic solubilizer.

3. A composition according to claim 2, wherein the amphipathic molecule is a lipid having a hydrophobic tail containing no more than 8 carbons.

4. A composition according to claim 1, wherein the at least one long-chain phospholipid is a phosphatidylcholine.

5. A composition according to claim 1, wherein the activity of the polymerase is enhanced in the presence of the solubilized phospholipid, capable of being measured in a replication assay by incorporation of deoxynucleoside triphosphate.

6. A composition according to claim 5, wherein the enhancement is at least 3-fold.

9. The composition of claim 1, wherein said thermostable polymerase is a Taq polymerase.

10. The composition of claim 1, wherein the phospholipid has at least one hydrophobic tail containing 8 to 20 carbons.

7. A kit, comprising:

a thermostable polymerase in a buffer containing a solubilized phospholipid having 8 to 30 carbons and a stabilizer.

8. A kit according to claim 7, wherein the stabilizer is a lipid having a hydrophobic tail containing no more than 8 carbons.