GSM8403746: MN-2,Normal,hepatocellular carcinoma; Homo sapiens; RNA-Seq
RNA extraction: Total RNA was extracted using the TRIzol method. The purity (OD260/280), concentration, and nucleic acid absorption peak of the RNA were assessed using a NanoPhotometer® spectrophotometer (IMPLEN, CA, USA). RNA integrity was evaluated using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system Sequencing libraries were generated using the NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA). mRNA was enriched using oligo(dT) magnetic beads. Fragmentation buffer was then added to break the mRNA into short fragments. Using these mRNA fragments as templates, first-strand cDNA was synthesized using random hexamer primers. Second-strand cDNA synthesis was performed using DNA Polymerase I, RNase H, dNTPs, and buffer. The cDNA fragments were purified with the AMPure XP system (Beckman Coulter, Beverly, USA) to select cDNA fragments of 150-200 bp in length, followed by polymerase chain reaction (PCR) to generate the final cDNA library.