title: GSM7770148: Stationary phase cell total replicate2; Saccharomyces cerevisiae; OTHER
library_name: GSM7770148
library_construction_protocol: exponential and stationary phase total RNA was extracted according to previous report (Liu et al., 2017). Polysome RNA were extracted with Trizol (Invitrogen, 10296010) and digested with DNase I (Ambion, AM2222) according to the manufacturer's recommendations. 2 ug total RNA was fragmented by heating at 95 °C for 40 mins. The first strand cDNA was transcribed by SuperScript III Reverse Transcriptase (Invitrogen, 18080044) with an T7 primer (AAGCTTAGATATCTAATACGACTCACTATAGGGACCTACACGACGCTCTTCCGATC (T) 20VN) tagged with Illumina A adaptor and T7 promoter, and then the second strand was synthesized with PrimeScript™ Double Strand cDNA Synthesis Kit (Takara, 6111A). The libraries were amplified by in vitro transcription with RiboMAX Large Scale RNA production Systems- T7 (Promega, P1320). Next, first strand cDNA was reversed with random primers (AGTTCAGACGTGCTCTTCCGATCTNNNNNN), and then cDNAs used as template to amplify the library. The P5 and P7 adapters were linked by PCR with Platinum Taq DNA polymerase (Invitrogen, 15966025) for 12-15 cycles. Finally, 200-500 bp library products were enriched by size selection with AMPure XP Beads (Beckman Coulter, A63881).