GSM6709836
Total RNA was isolated using a RiboPure RNA Purification Kit (Thermo Fisher). Kit buffers were supplemented with 10mM BME. Nascent RNAs were purified using MTS-biotin and streptavidin beads as described in Duffy et al. 2015. Nascent RNA barcodes were reverse transcribed with a reporter-specific primer. PCR1 was performed with primers complementary to the RT product and containing overhanging regions for multiplexing PCR. qPCR was used to determine the number of amplification cycles for PCR2. PCR2 was perfomed using unique dual indexes.