SD2
Oligo(dT)-attached magnetic beads were used to purified mRNA. Purified mRNA was fragmented_x000D_ into small pieces with fragment buffer at appropriate temperature. Then First-strand cDNA was_x000D_ generated using random hexamer-primed reverse transcription, followed by a second-strand cDNA_x000D_ synthesis. afterwards, A-Tailing Mix and RNA Index Adapters were added by incubating to end_x000D_ repair. The cDNA fragments obtained from previous step were amplified by PCR, and products_x000D_ were purified by Ampure XP Beads, then dissolved in EB solution. The product was validated on_x000D_ the Agilent Technologies 2100 bioanalyzer for quality control. The double stranded PCR products_x000D_ from previous step were heated denatured and circularized by the splint oligo sequence to get the_x000D_ final library. The single strand circle DNA (ssCir DNA) was formatted as the final library. The_x000D_ final library was amplified with phi29 to make DNA nanoball (DNB) which had more than 300_x000D_ copies of one molecular, DNBs were loaded into the patterned nanoarray and single end 50 bases_x000D_ reads were generated on BGIseq610 platform (BGI-Shenzhen, China).
SUB14994853
111