Methods and kits for joining fragmented nucleic acids together
AssigneeRoche Molecular Systems, Inc.
InventorsStephen G. Will, Yuker Wang
Life of this patent
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Abstract
Methods and kits for joining fragmented nucleic acid sequences together are provided, including performing an amplifying step including contacting a sample suspected of including a fragmented target nucleic acid with a pair of external primers and a pair of self-complementary internal primers, and generating a full length target nucleic acid. The methods can include performing an amplifying step, a hybridizing step, and a detecting step. Furthermore, kits are provided that are designed for the detection of a target nucleic acid sequence.
Claims17
A method for joining fragmented nucleic acid together, comprising the steps of:
- performing an amplifying step comprising contacting a sample suspected of comprising a fragmented target nucleic acid with:
- a pair of external primers comprising a first nucleic acid sequence and a second nucleic acid sequence to produce a first amplification product comprising a first sense strand and a first anti sense strand if any target nucleic acid is present in the sample;
- and
- a pair of self-complementary internal primers comprising a third nucleic acid sequence and a fourth nucleic acid sequence, the third nucleic acid sequence configured to hybridize to the first antisense strand of the first amplification product to produce a second sense strand of a second amplification product and the fourth nucleic acid sequence configured to hybridize to the first sense strand of the first amplification product to produce a second anti-sense strand of the second amplification product;
- and
- generating a full length target nucleic acid by:
- annealing a first 3′ end region of the first sense strand with a second 3′ end region of the first anti-sense strand, wherein the first 3′ end region of the first sense strand primes the extension of the first sense strand over the first anti-sense strand, and the second 3′ end region of the first anti-sense strand primes the extension of the first anti-sense strand over the first sense strand;
- and
- annealing a third 3′ end region of the second sense strand with a fourth 3′ end region of the second anti-sense strand, wherein the third 3′ end region of the second sense strand primes the extension of the second sense strand over the second anti-sense strand, and the fourth 3′ end region of the second anti-sense strand primes the extension of the second anti-sense strand over the second sense strand.
A kit for joining fragmented nucleic acid together, comprising:
- a pair of external primers comprising a first nucleic acid sequence and a second nucleic acid sequence to produce a first amplification product comprising a first sense strand and a first anti-sense strand if any of the target nucleic acid is present in the sample;
- and
- a pair of self-complementary internal primers comprising a third nucleic acid sequence and a fourth nucleic acid sequence, the third nucleic acid sequence configured to hybridize to the first antisense strand of the first amplification product to produce a second sense strand of a second amplification product and the fourth nucleic acid sequence configured to hybridize to the first sense strand of the first amplification product to produce a second anti-sense strand of the second amplification product.
Dependencies are inferred from the claim text.