GSM3757989: MVm_miRNA-Seq; Valsa mali; miRNA-Seq
According to the instruction of TrizolTM Reagent (Invitrogen, Carlsbad, CA), total RNA was extracted from each sample, and RNA purity was evaluated using NanoPhotometer® spectrophotometer (IMPLEN, CA, USA). RNA Nano 6000 Assay Kit and Agilent Bioanalyzer 2100 system (Agilent Technologies, CA, USA) were used to assess RNA concentration and integrity. Three repeats of control samples and inoculation samples were mixed for small RNA library construction, respectively. Three μg of total RNA per sample was used as input material for the small RNA library. Three micrograms total RNA was used for each small RNA library. Sequencing libraries were generated using NEBNext® Multiplex Small RNA Library Prep Set for Illumina® (New England BioLabs Inc, Beverly, MA, USA) according to manufacturer's instructions. The clustering of each index-coded samples was performed on a cBot Cluster Generation System using TruSeq SR Cluster Kit v3-cBot-HS (Illumia, San Diego, USA) following the manufacturer's instructions.  Approximately 20 μg total RNA used for small RNA sequencing library construction were also used to construct degradome sequencing library following protocol as described (German et al., 2009). Poly(A)+ RNA was isolated and annealed with biotinylated random primers. 5' adaptor was ligated to the annealed poly(A)+ RNAs which contain 5'-monophosphates. After reverse transcription and PCR amplification, single-end sequencing (36 bp) were performed on Illumina Hiseq2500 at the LC-BIO (Hangzhou, China) following manufacturer's protocol.
GEO Accession
GSM3757989