title: GSM8033253: Gro-α; Homo sapiens; RNA-Seq
library_construction_protocol: TRIrizol reagent was used to purify the total RNA of isolated circulating neutrophils according to the manufacturer's instruction. After qualifying the RNA samples, mRNA is enriched using magnetic beads with Oligo(dT). Then, the mRNA is broken into short fragments by adding fragmentation buffer. Use the mRNA as a template to synthesize first-strand cDNA using random hexamers. Next, add buffer, dNTPs, DNA polymerase I, and RNase H to synthesize second-strand cDNA. Double-stranded cDNA is then purified by AMPure XP beads. Repair the ends of the purified double-stranded cDNA, and ligate sequencing adapter by adding A-tails. Perform size selection of the cDNA fragments using AMPure XP beads. Finally, perform PCR amplification, purify the PCR products using AMPure XP beads, and obtain the final library. The constructed library is quantitated using Qubit 2.0. Agilent 2100 is used to detect the size of the inserted fragments then. Once the inserted fragments meet the expected size, qPCR is finally employed to accurately quantify the effective concentration of the library to ensure its quality.