GSM5050764: Retina Ptf1a-/- rep2; Mus musculus; miRNA-Seq
Total retina RNA was extracted using TRIzol, then denatured and separated on a 15% TBE urea gel. 18-30 nt small RNA was recovered from a gel slice. Gel purified small RNA from 1 µg total RNA was used to prepare a small RNA library using a New England Biolabs Next Multiplex Small RNA Library Prep Kit according the manufacturer's directions, with the following modification: after reverse transcription, the reaction was denatured, separated on a 10% TBE urea gel; a gel slice corresponding to 60 to 80 nt was isolated (based on size markers in an adjacent lane: low range ssRNA ladder, NEB). The reverse transcribed product in the gel slice was eluted, precipitated by adding 2-propanol, suspended in Tris-Hcl buffer, and then used in subsequent steps.