title: GSM8544686: Ezh2a, wild type, rep3; Mus musculus; RNA-Seq
library_construction_protocol: Total RNA was isolated using Trizol Reagent (Invitrogen Life Technologies). The quality and quantity of the total RNA were assessed using a NanoDrop spectrophotometer (Thermo Scientific) and Qubit 4.0 for purity and concentration evaluation, while RNA integrity was measured using the Agilent 2100/4200 system. Library construction involved the purification of mRNA from three micrograms of total RNA using poly-T oligo-attached magnetic beads. Following fragmentation with divalent cations, the first strand of complementary DNA (cDNA) was synthesized using random hexamers, followed by second strand synthesis with buffer, dNTPs, RNase H, and DNA polymerase I. The double-stranded cDNA was repaired and adenylated at the 3' end. Subsequent purification and fragment selection were performed using Hieff NGS® DNA Selection Beads, and the library was amplified via PCR and quantified with Qubit. The target region library was denatured and processed to generate single-stranded circular DNA, which was amplified through rolling circle amplification (RCA) to produce DNA nano balls (DNB).