GSM7027461: CAGE-seq_BT549 cells, VEH treatment, BioRep2; Homo sapiens; RNA-Seq
CAGE-seq analysis performed following a previously described protocol (Cvetesic et al., 2018). In brief, sample RNA was mixed with carrier RNA followed by first-strand cDNA synthesis. The ends of the RNA were oxidized, biotinylated and RNA fragments not overlapping first-strand cDNA were RNase digested. This provided a library of fragments where the biotinylated 5'cap was pulled down using streptavidin paramagnetic beads. The cap-trapped cDNA strand was then released by RNase treatment and the linker was ligated to the first cDNA strand followed by second-strand cDNA synthesis. The carrier was degraded, and the libraries were then purified using SPRI paramagnetic beads, PCR amplified and subjected to sequencing on Illumina NextSeq 550 to a minimum of 35M reads per sample.