title: GSM8369929: HCT-116-shTfR1-CDDP-RNA-seq 3; Homo sapiens; RNA-Seq
library_construction_protocol: HCT-116 cells transfected with shSCR or shTFRC were seeded in 100 mm dishes overnight. Cells were harvested after CDDP treatment and washed with PBS. TRIzolTM reagent (Thermo Scientific, 15596026) was used to extract total RNA from each sample. After passing the detection of total RNA samples, purification is carried out based on the characteristics of mRNA from different sources. The end of eukaryotic mRNA3 has a polyA tail structure, and magnetic beads with Oligo (dT) were selected for enrichment and purification; Prokaryotic mRNA does not possess this characteristic, so a reagent kit was used to remove rRNA to obtain more effective information. Add Fragmentation Buffer to the purified mRNA to make its fragments short. Using the fragmented RNA as a template, reverse transcription was performed using random primers to synthesize the first strand of cDNA, and the second strand of cDNA was synthesized by adding the 2nd Strand Marking Buffer and 2nd Strand/End Repair Enzyme Mix. After end repair, addition of base A, sequencing adapter, magnetic bead screening to recover the target fragment, and PCR amplification, the entire library preparation work was completed.