RNA-seq CM Mrg15 WT-2
1 g total RNA was used for library preparation. Poly(A) mRNA isolation was performed using Oligo(dT) beads (Vazyme, Cat. No. N401). RNA-seq libraries were prepared using the VAHTS Universal V8 RNA-seq Library Prep Kit for Illumina (Vazyme, Cat. No. NR605-02) according to the manufacturers protocol. Briefly, mRNA fragmentation was achieved using divalent cations and high temperature, followed by priming with Random Primers. First-strand cDNA and then second- strand cDNA were synthesized. The purified double-stranded cDNA was treated to repair both ends and was subjected to dA-tailing in a single reaction, followed by T-A ligation to add adaptors to both ends. Size selection of adaptor-ligated DNA was performed using DNA Clean Beads. Each sample was then amplified by PCR using P5 and P7 primers, and the PCR products were validated. Libraries with different indices were multiplexed and loaded onto an Illumina NovaSeq instrument for sequencing using a 150 paired-end configuration according to manufacturers instructions.