GSM8428964: PROseq_Myxo_L2hgdh_Rep1; Mus musculus; OTHER
performed according to the qPRO-seq protocol (Judd, J. et al. https://doi.org/10.1101/2020.05.18.102277). Briefly, 2.5 million mESC nuclei and 1% spike-in fly S2 nuclei were used for nuclear run-on assay with biotin-11-UTP/CTP/ATP/CTP, 40 µM each (Revvity). REV3 adaptor was ligated to 3' end of fragmented nuclear RNA using T4 RNA ligase 1 (NEB). 5' hydroxyl repair with PNK (NEB), decapping with RppH (NEB), and REV5 ligation to 5' end of RNA were performed on Dynabeads MyOne Streptavidin C1 beads (ThermoFisher). cDNA was generated using SuperScript III (ThermoFisher). Libraries were amplified using Phusion Hot Start II (ThermoFisher) with 15 PCR cycles and sequenced on NextSeq 2000 (Illumina)