Compositions and methods for negative selection of non-desired nucleic acid sequences

A method of creating a strand retained library of nucleic acids, the method comprising:

• a) annealing a partial duplex primer to a nucleic acid template strand, wherein the nucleic acid template strand comprises a non-canonical nucleotide, wherein the partial duplex primer comprises a 3′ overhang;
• b) extending the partial duplex primer to form a primer extension product strand, thereby forming a double-stranded nucleic acid comprising (i) the primer extension product strand and (ii) the nucleic acid template strand;
• c) ligating an adaptor to a 5′ end of the primer extension product strand;
• and
• d) cleaving the nucleic acid template strand from the double-stranded nucleic acid, wherein the cleaving comprises use of a cleavage agent specific for the non-canonical nucleotide, thereby creating a strand retained library of nucleic acids.

A method of depleting or reducing a non-desired nucleic acid sequence from a nucleic acid library comprising the non-desired nucleic acid sequence in a non-desired DNA fragment having an adaptor at each end and a desired nucleic acid sequence in a desired DNA fragment having an adaptor at each end, the method comprising:

• a) annealing a sequence-specific oligonucleotide probe to the non-desired nucleic acid sequence in the non-desired DNA fragment having an adaptor at each end, wherein a 3′ end of the sequence-specific oligonucleotide probe comprises perfect complementarity to a portion of the non-desired nucleic acid sequence, thereby producing a DNA fragment with a partial duplex;
• b) treating the DNA fragment with the partial duplex with a nuclease that cleaves at a double-stranded site in the partial duplex;
• and
• c) amplifying the desired nucleic acid sequence in the desired DNA fragment having an adaptor at each end by performing polymerase chain reaction (PCR) with a set of primers that anneal to the adaptor at each end of the desired DNA fragment, thereby generating amplified desired DNA fragments.