Methods for analysis of molecular events
First Claim
Patent Images
1. A method for detecting a variant in a primer extension assay, the method comprising the steps of:
- a) contacting a target nucleic acid with a nucleic acid primer complementary to a region of said target nucleic acid; and
b) extending said primer in the presence of;
i) a first nucleotide that is complementary to a first variant nucleotide suspected to be at a position downstream of said region; and
, ii) a second nucleotide that is complementary to a second variant nucleotide at said position, thereby to reduce misincorporation of said first nucleotide on a template comprising said second variant nucleotide..
3 Assignments
0 Petitions
Accused Products
Abstract
Methods and compositions are provided for detecting the presence of nucleic acid sequence variants in a subpopulation of nucleic acid molecules in a biological sample. These methods are particularly useful for identifying individuals with mutations indicative of cancer.
-
Citations
33 Claims
-
1. A method for detecting a variant in a primer extension assay, the method comprising the steps of:
-
a) contacting a target nucleic acid with a nucleic acid primer complementary to a region of said target nucleic acid; and
b) extending said primer in the presence of;
i) a first nucleotide that is complementary to a first variant nucleotide suspected to be at a position downstream of said region; and
,ii) a second nucleotide that is complementary to a second variant nucleotide at said position, thereby to reduce misincorporation of said first nucleotide on a template comprising said second variant nucleotide.. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15)
-
-
16. A method for minimizing nucleotide misincorporation in a primer extension assay, the method comprising the steps of:
-
a) contacting a target nucleic acid with a nucleic acid primer complementary to a region of said target nucleic acid; and
b) extending said primer in the presence of;
i) a first nucleotide that is complementary to a first variant base suspected to be at a position downstream of said region, and a first polymerase that preferentially incorporates said first nucleotide; and
,ii) a second nucleotide that is complementary to a second variant base at said position and a second polymerase that preferentially incorporates said second nucleotide, thereby to reduce misincorporation of said first nucleotide in a primer extension reaction at said second variant base. - View Dependent Claims (17, 18, 27, 30, 32, 33)
-
-
19. A method for detecting the presence of a variation at a predetermined position in a heterogeneous nucleic acid sample, the method comprising the steps of:
-
a) contacting a target nucleic acid with a nucleic acid primer substantially complementary to said target nucleic acid to form a nucleic acid complex upstream from a predetermined position in said target nucleic acid; and
b) contacting said nucleic acid complex with a first nucleotide, a first polymerase, a second nucleotide, and a second polymerase, wherein said first nucleotide is complementary to a first variant base at said predetermined position, and said second nucleotide is complementary to a second variant base at said predetermined position, and wherein said first and second polymerases are different, thereby to reduce misincorporation of said first nucleotide in a primer extension reaction at a said second variant base. - View Dependent Claims (20, 21, 22, 23, 24, 25, 26, 28, 29)
-
-
31. A kit comprising:
-
a) a first nucleotide, a second nucleotide; and
b) a first polymerase adapted for said first nucleotide and a second polymerase adapted for said second nucleotide.
-
Specification