Patent ID: 9708652
Date: 2017-07-18
CPC Classifications: C12N,C12Q

Claim:
1. A method of preparing a DNA molecule, comprising the steps of: (1) providing a DNA molecule; (2) providing an oligonucleotide-linked DNA molecule in a single incubation, said single incubation comprising: (a) modifying the ends of the DNA molecule to provide attachable ends; (b) ligating a first oligonucleotide comprising a known sequence and a nonblocked 3, end to an end of the DNA molecule to produce an oligonucleotide-linked molecule, wherein the 5′ end of the DNA molecule is attached to the nonblocked 3, end of the first oligonucleotide, leaving a nick site between a juxtaposed 3′ end of the DNA molecule and a 5′ end of the first oligonucleotide; and (c) extending the juxtaposed 3, end of the DNA molecule from the nick site by polymerization; (3) digesting the oligonucleotide-linked molecule with a mixture of methylation-sensitive restriction enzymes that do not cleave within the first oligonucleotide of the oligonucleotide-linked molecule; and (4) amplifying the digested oligonucleotide-linked molecule with a primer complementary to at least a portion of the known sequence of the first oligonucleotide to produce amplified oligonucleotide-linked molecules.