2.6 RNA-Seq Raw Data Clean and Alignment
Raw reads covering more than 2-N bases were first cast off. Then adaptors and low-quality bases were removed from raw sequencing reads using FASTX-Toolkit (Version 0.0.13). The short reads less than 16nt were also discarded. Then, clean reads were aligned to the GRch38 genome by tophat2 [28] allowing 4 mismatches. Uniquely mapped reads were utilized for gene reads number counting and FPKM calculation (fragments per kilobase of transcript per million fragments mapped) [29].