RNA-seq of tumor  cell
Oligo(dT)-attached magnetic beads were used to purified mRNA.  Purified mRNA was fragmented  into small pieces with fragment buffer at appropriate temperature.  Then First-strand cDNA was  generated using random hexamer-primed reverse transcription, followed by a second-strand cDNA  synthesis.  afterwards, A-Tailing Mix and RNA Index Adapters were added by incubating to end  repair.  The cDNA fragments obtained from previous step were amplified by PCR, and products  were purified by Ampure XP Beads, then dissolved in EB solution.  The product was validated on  the Agilent Technologies 2100 bioanalyzer for quality control.  The double stranded PCR products  from previous step were heated denatured and circularized by the splint oligo sequence to get the  final library.  The single strand circle DNA (ssCir DNA) was formatted as the final library.  The  final library was amplified with phi29 to make DNA nanoball (DNB) which had more than 300  copies of one molecular, DNBs were loaded into the patterned nanoarray and pair end 100 bases  reads were generated on MGIseq2000 platform.