Identification of edited miRNAs in glioblastoma

MiRNA capturing and library construction were conducted using Illumina's TruSeq Small RNA Sample Prep Kit following the manufacture protocol. Briefly, 3' adapter was ligated to total RNA using truncated T4 RNA Ligase 2 without ATP, then 5' adapter was added using T4 RNA Ligase with ATP. Afterwards, reverse transcription followed by PCR was used to create cDNA constructs based on the small RNA ligated with 3' and 5' adapters. The introduction of index sequence (bar-codes) was done at the PCR step. Finally, gel electrophoresis was used to purify the amplified cDNA construct in preparation for subsequent cluster generation. After library quality check by Bioanalyzer High Sensitivity chip, clusters were generated and libraries were run on HiSeq2000 by using standard Illumina sequencing workflow with the multiplexing option. The mature miRNA libraries from the A172 cell-line were sequenced on two lanes of Illumina HiSeq2000 instrument following the manufacture protocol. In each sequencing lane two bar-codes were used, one for each tested condition (siADAR2 and control). Primary data analyses of sequence data and quality controls were performed using the Illumina pipeline (version 1.7). The reads were filtered using Illumina's HiSeq2000 softwares, Illumina indexes separation was also performed.