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US 2014/0235845 A1Published application
Sequence amplification with loopable primers
AssigneeLife Technologies Corporation
InventorsKAI LAO, NEIL STRAUS
Life of this patent
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Abstract
The present disclosure relates to the amplification of target nucleic acid sequences. This can be accomplished via the use of various primers. The use of these primers, as described herein, results in nucleic acid structures that can reduce the amplification of nonspecific hybridization events (such as primer dimerization) while allowing the amplification of the target nucleic acid sequences.
Claims19
A loopable primer comprising:
- a 3′ target specific region;
- a first loop forming region;
- a universal region;
- and
- a second loop forming region,
- wherein the first loop forming region and the second loop forming region are configured to hybridize to each other to form a stem and a loop,
- wherein the universal region is located between the first and second loop forming regions,
- wherein the stem and the loop, when formed, are configured to allow the 3′ target specific region to hybridize to a target nucleic acid without the remainder of the loopable primer annealing to the target nucleic acid, and
- wherein the universal region is configured to form a self-hybridized structure on subsequent amplifications.
6 more elements
The self-hybridizable DNA structure in a PCR amplification mixture, said self-hybridizable DNA sequence comprising:
- a first loop forming region;
- a universal region connected to the first loop forming region;
- a second loop forming region connected to the universal region,
- wherein the first loop forming region and the second loop forming region are complementary to each other;
- a first 3′ target specific region connected to the second loop forming region;
- a sequence that is complementary to a second 3′ target specific region, wherein the sequence that is complementary to the second 3′ target specific region is part of a same nucleic strand as the first 3′ target specific region;
- a sequence that is complementary to the second loop forming region, wherein the sequence that is complementary to the second loop forming region is connected to the sequence that is complementary to the second 3′ target specific region;
- a sequence that is complementary to the universal region, wherein the sequence that is complementary to the universal region is connected to the sequence that is complementary to the second loop forming region;
- and
- a sequence that is complementary to the first loop forming region, wherein the sequence that is complementary to the first loop forming region is connected to the sequence that is complementary to the universal region,
- wherein the self-hybridizable DNA structure is configured to form a stem and a loop structure, wherein the stem is formed by hybridization of the sequence comprising the first loop forming region, the universal region and the second loop forming region to the comprising their respective complementary sequences.
8 more elements
15-45. (canceled)
A loopable primer comprising:
- a 3′ target specific region;
- a first loop forming region;
- a universal region;
- a noncomplementary region;
- and
- a second loop forming region,
- wherein the first loop forming region and the second loop forming region are configured to hybridize to each other to form a stem and a loop,
- wherein the universal region is located between the first and second loop forming regions, wherein the noncomplementary region is located between the first and second loop forming regions,
- wherein the stem and the loop, when formed, are configured to allow the 3′ target specific region to hybridize to a target nucleic acid without the remainder of the loopable primer annealing to the target nucleic acid, and
- wherein the universal region is configured so that a sequence associated with a target sequence that has been amplified by the loopable primer can form a self-hybridized structure comprising the universal region, the noncomplementary region, the first loop forming region, and the second loop forming region on subsequent amplifications, wherein said loopable primer is created by deliberately selecting the sequence of the first loop forming region so that it hybridizes to the second loop forming region, and wherein said loopable primer is created by deliberately selecting a universal region.
7 more elements
Dependencies are inferred from the claim text.