Provided herein is a method of preparing an RNA sample comprising: a) obtaining an RNA sample comprising: i. long RNA molecules that may be unfragmented or fragmented to contain 5′-OH group and a 2′-3′-cyclic phosphate group; and ii. short RNA molecules that comprise a 5′ phosphate group and a 3′ OH group; and b) contacting the RNA sample with an adaptor comprising either a 2′-PO group and 3′-OH group or a 2′,3′-cyclic phosphate group in the presence of a eukaryotic tRNA ligase, thereby producing a ligated RNA sample in which a) the short RNA molecules are selectively ligated to the adaptor or b) the short RNA molecules and long RNA fragments are selectively ligated to the adaptor.