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Several recent reviews expertly address the relative merits of different approaches to preparation and analysis of deep-sequenced small RNA libraries. Here, we focus on an array of protocols and tools with the intention of assisting researchers in improving short RNA profiles constructed with second-generation sequencing. This includes methods and commentaries on the preparation of sequencing-caliber immunoprecipitation RNA libraries, techniques for targeting different populations of RNAs with distinct 5′- and 3′-ends, reduction of adapter dimers in libraries, and dealing with the underappreciated problem of genomic cross-mapping of similar miRNA sequences. |