Methods and kits for depleting undesired nucleic acids

Abstract

Methods and kits for depleting amplicons that correspond to undesired RNA species present in a sample are provided. The disclosed methods and kits employ a blocker that anneals with at least a portion of the undesired RNA, resulting in a duplex that is not a suitable substrate for ligating an adapter to the end of the undesired RNA.

1 . A kit comprising: at least one single-stranded nucleic acid blocker complementary to at least a portion of an undesired RNA species, wherein the single-stranded nucleic acid blocker does not form a unimolecular duplex under conditions suitable for the single-stranded nucleic acid blocker to anneal with the undesired RNA to form an RNA-blocker duplex; at least one 3′ adapter, and at least one 5′ adapter. 
     
     
         2 . The kit of  claim 1 , wherein at least one 3′ adapter comprises an rApp at its 5′ end, a dideoxy nucleotide at its 3′ end, or an rApp at its 5′ end and a dideoxy nucleotide at its 3′ end. 
     
     
         3 . The kit of  claim 1 , further comprising at least one ligase. 
     
     
         4 . The kit of  claim 1 , wherein the ligase comprises at least one of: T4 RNA ligase 2, truncated T4 RNA ligase 2, T4 RNA ligase 1, or  Methanobacterium  thermoautotrophicum RNA ligase. 
     
     
         5 . The kit of  claim 1 , wherein the blocker comprises at least one deoxyribonucleotide, at least one ribonucleotide, at least one LNA, at least one 2′ o-methyl nucleotide, at least one deoxyuridine, at least one inosine, or combinations thereof.