GSM5697075: RNAseq_C2_H; Mus musculus; RNA-Seq
Total RNA was extracted by ZYMO Research Quick-RNA MiniPrep kit from lysed 10 million of cells and spheroids in RNA lysis buffer, then removed most of gDNA with the spin-away filter. After that, the mixture of RNA was transferred with ethanol to Zymo-Spin IIICG column to remove trace DNA by DNase I on the column, then washed twice with RNA wash buffer followed by elution with 50 μl DNase/RNase-free water. RNA-seq library was prepared with NEBNext® Poly(A) mRNA Magnetic Isolation Module (NEB #E7490). The Oligo dT Beads were washed with RNA binding buffer and incubated with a total 1 μg RNA to purify mRNA, followed with more washing by beads washing buffer. Then mRNA was eluted with elution buffer and reverse transcribed. After that, the first and the second strand cDNA were synthesized. After purification of Double-stranded DNA, an adaptor was added. Adaptor-ligated DNA was enriched by PCR followed by purification, then the DNA library was sequenced with Illumina HiSeq3000.