GSM8878611: Tumor tissue, PBS-treated, rep2; Mus musculus; RNA-Seq
The tumor was removed and frozen in liquid nitrogen.The tissue sample were homogenized the lysated using a rotor-stator homogenizer in Trizol. After DNA removal by AllPrep DNA spin column,the RNA were collected using a RNeasy spin column. After two washes, the RNA was dissolved in RNase-free water. Fragmented mRNA was used as a template to synthesize the first-strand cDNA using random hexamer primers and M-MuLV reverse transcriptase. The RNA strand was then degraded with RNase H, and the second-strand cDNA was synthesized using DNA polymerase I and dNTPs. The resulting double-stranded cDNA was purified, followed by end repair, A-tailing, and ligation of sequencing adapters. Fragments of approximately 250–300 bp were selected using AMPure XP beads. The selected fragments were PCR-amplified and purified again with AMPure XP beads to obtain the final sequencing library.