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US 9,175,325 B2Granted
Global amplification using a randomly primed composite primer
AssigneeNuGEN Technologies, Inc.
InventorsNurith Kurn, Shenglong Wang
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Abstract
The invention relates to the field of polynucleotide amplification. More particularly, the invention provides methods, compositions and kits for amplification of (i.e., making multiple copies of) a multiplicity of different polynucleotide template sequences using a randomly primed RNA/DNA composite primer.
Claims17
A method for amplification of a template DNA, comprising:
- (a) incubating the template DNA with
- (i) a pool of first composite primers, each first composite primer comprising
- (1) a 5′ RNA portion that is not hybridizable to the template DNA and
- (2) a 3′ DNA portion, wherein the 3′ DNA portion of each of the first composite primers comprises a random sequence that is different from the 3′ DNA portion of the other first composite primers in the pool, and
- (ii) a DNA polymerase having strand displacement activity, whereby a complex comprising an RNA/DNA heteroduplex is generated;
- (b) incubating the complex under conditions that permit cleavage of RNA in the RNA/DNA heteroduplex;
- and
- (c) incubating the complex with an amplification primer, wherein the amplification primer is a composite primer comprising a 5′ RNA portion and a 3′ DNA portion, wherein the amplification primer anneals to sequence complementary to the RNA sequence cleaved from the RNA/DNA heteroduplex, whereby multiple copies of an amplification product are generated.
5 more elements
Dependencies are inferred from the claim text.