GSM2845334: techrep_A+_3h.2; Danio rerio; OTHER
Total RNA was isolated using TRIzol (Invitrogen), after adding 120fg of mRNA with 5 known 3'UTR control sequences into each RNA sample during the initial TRIzol lysis step. In the first step, total-RNA was reverse-transcribed with Maxima RT (Thermo Fisher) and a gene-specific primer that matched the constant 3'UTR of mRNA reporters. RT primer also added a random 8nt UMI and a constant RT adaptor sequence. In the second step, resulting cDNA was amplified by 18 cycles of Phusion PCR with primers that matched the reporter's constant 3'UTR sequence and the RT adaptor. PCR primers also added the appropriate Illumina sample barcodes and sequencing. adaptors