GSM8795354: SCC1_KO_2_BX_795_Rep3; Homo sapiens; RNA-Seq
Total RNA was isolated using TRIzol reagent (Sigma), and its quality was assessed with the NanoDrop Spectrometer (ND-1000 Spectrophotometer, Peqlab). High-quality RNA samples (20 g each) were dispatched to the Beijing Genomics Institute (BGI, Shenzhen) for cDNA library construction and sequencing. DNase I treatment was applied to the total RNA samples to eliminate any potential DNA contamination. Following this, mRNA was selectively enriched using oligo (dT) magnetic beads, and then the mRNA was fragmented to approximately 200 bp. The first strand of cDNA was synthesized with random hexamer primers, and subsequently, a mixture of buffer, dNTPs, RNase H, and DNA polymerase I was added to facilitate the synthesis of the second strand. The resulting double-stranded cDNA was purified using magnetic beads. Next, end repair and the addition of a single adenine nucleotide at the 3' end were performed. Sequencing adapters were then ligated to the cDNA fragments, which were subsequently enriched through PCR amplification. During the quality control phase, the sample library was qualified and quantified using the Agilent 2100 Bioanalyzer and the ABI Step-One-Plus Real-Time PCR System. The library was loaded onto the Illumina HiSeq™ 2000 platform for sequencing.