In RNA-seq, highly expressed transcripts with minimal biological interest, such as ribosomal RNA (rRNA) can dominate readouts and mask detection of more informative low-abundance transcripts. This challenge is amplified when working with sample types for which pre-designed RNA depletion kits are not available. The NEBNext RNA Depletion Core Reagent set provides a customized approach to deplete unwanted RNA from any organism, using probe sequences designed with the user-friendly NEBNext Custom RNA Depletion Design Tool. The efficient RNase-H-based workflow, and close tiling of probes designed using the online tool, enables effective depletion from both low- and high-quality RNA, with a broad range of input amounts. The kit is also available with RNAClean beads.