GSM4563830: HC_2; Homo sapiens; RNA-Seq
Neutrophils were isolated by density gradient centrifugation on Polymorphprep (Axis-Shield, Dundee, UK) according to the manufacturer's instructions. Oligo(dT)-attached magnetic beads-purified mRNA was fragmented into pieces at appropriate temperature. cDNA was generated by random hexamer-primed reverse transcription. Afterwards, RNA Index Adapters and A-Tailing Mix were added to end repair. The cDNA fragments were amplified by polymerase chain reaction (PCR), and products were purified using Ampure XP Beads. The products were qualified on Agilent Technologies 2100 bioanalyzer. The double-strand PCR products were denatured and circularized by the splint oligo sequence to construct the final library. Then the single-strand circle DNA was formatted as the final library. The final library was amplified by phi29 to produce DNA nanoball (DNB) with more than 300 copies of one molecule. DNBs were loaded into patterned nanoarray and single end 50 bases reads were generated on BGIseq500 platform (BGI-Shenzhen, China).