Libraries were prepared from total RNA using a SMARTer Stranded RNA-Seq Kit (Clontech Laboratories, Inc., USA). mRNA was isolated with Poly (A) RNA Selection Kit (LEXOGEN, Inc., Austria); indexed with Illumina indices 1-12; and enriched with PCR. Libraries were assessed for mean fragment size using an Agilent 2100 Bioanalyzer (DNA High Sensitivity Kit), quantified with StepOne Real-Time PCR System (Life Technologies, Inc., USA). High-throughput sequencing was made as paired-end 100 sequencing with HiSeq 2500 system (Illumina, Inc., USA). FastQC (https://www.bioinformatics.babraham.ac.uk/projects/fastqc/ ) for quality control of raw sequencing data. Adapters, and low-quality reads (http://hannonlab.cshl.edu/fastx_toolkit/) and BBMap (https://sourceforge.net/projects/bbmap/ ), trimmed reads were mapped to reference with TopHat.55 Gene expression levels were estimated based on RC (read count) and FPKM (fragments per kb per million reads) values determined using BEDTools56 and Cufflinks.57 (link) then normalized with EdgeR within R (https://www.r-project.org ) Quantile normalization. Data mining and graphic visualized with ExDEGA (E-Biogen, Inc., Seoul, Republic of Korea). RNA-sequencing data have been stored at Gene Expression Omnibus database (www.ncbi.nlm.nih.gov/geo accession no. GSE183220).
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