RNA-Seq_PNT_8h_NC_ActD (CRX1623092)
Before RNA-seq library construction, add 0.1 uL of ERCC spike-in RNA control to each 1 ug RNA sample.The VAHTS Total RNA-seq (H/M/R) Library Prep Kit for Illumina (VAHTS, Catalog No. NR603) was used for RNA library preparation. In brief, 1 ug total RNAs were hybridized with the rRNA probe (H/M/R) and digested by RNase H to remove ribosomal RNAs. After DNase I digestion, the ribosomal-depleted RNAs were fragmented at 94 degrees Celsius for 8 min. Then the first-strand and second-strand cDNAs were synthesized successively using the provided reagents. The cDNA was purified by VAHTS DNA Clean Beads (VAHTS, Catalog No. NR415), followed by end repaired, dA-tailing, adapter ligation, and second strand cDNA digestion. After two-round of purification, the cDNAs was PCR-amplified and purified by VAHTS DNA