GSM9630400: HaCaT VitD; Homo sapiens; RNA-Seq
A 100-μl aliquot of cell lysate was mixed with 700μl of QIAzol (Qiagen) and used for total RNA extraction. The remaining lysate was incubated on ice for 10min and centrifuged at 800g for 5min at 4 °C. The pellets were washed once with 200μl of Buffer A. The cDNAs were synthesized from nascent RNA using random primers. The ribose diols in the 5' cap structures of RNAs were oxidized, and then biotinylated. The biotinylated RNA/cDNAs were selected by streptavidin beads (cap-trapping). After RNA digestion by RNaseONE/H and adaptor ligation to both ends of cDNA, double-stranded cDNA libraries (CAGE libraries) were constructed.