Patent ID: 9453257
Filing Date: 2016-09-27
CPC Classification: C12N,C12Q

Claim Text:
1. A method for selectively amplifying cell-free target nucleic acid from a biological sample containing a mixture of the target nucleic acid and non-target nucleic acid and for specifically amplifying a nucleic acid region of interest in the target nucleic acid, comprising: a) mixing said biological sample with a 5′ adapter molecule and a 3′ adapter molecule, wherein: b) without subjecting the mixture of a) to a process that generates blunt ends, introducing a ligase to the mixture of a), whereby the 5′ adapter strand of the double-stranded adapter complex generated in a) ligates, at the blunt end of the complex, to the target nucleic acid fragments at each of the 5′ phosphorylated ends, thereby generating a ligated sample comprising double-stranded adapter complex ligated to the 5′ phosphorylated ends of the target nucleic acid fragments; c) heating the ligated sample of b) to release the 3′ adapter strands of the double-stranded adapter complex ligated to the 5′ phosphorylated ends of the target nucleic acid fragments, thereby generating products comprising single-stranded 5′ adapter strands comprising 5′ protruding ends at each end of the target nucleic acid; d) adding a polymerase to fill in the single-stranded 5′ protruding ends of the 5′ adapter strands in the products generated in c); e) adding 5′ adapter primers that hybridize to the filled in 5′ ends of the products of d) and amplifying the products of d) under conditions comprising short extension times of up to about 10 seconds, whereby the target nucleic acid is selectively amplified and enriched; and f) performing an additional amplification step after e), wherein the amplification is performed using a target-specific PCR primer and whereby the nucleic acid region of interest is specifically amplified in the target nucleic acid.