RNA-Seq of SMMC-7721
mRNA molecules were purified using poly-T-oligo-attached magnetic beads, and fragmented into small pieces using divalent cations under elevated temperature. The cleaved RNA fragments were used to synthesize the first strand cDNA using reverse transcriptase and random primers, followed by a second-strand cDNA. After the cDNA fragments were subjected to 3 adenylation and ligation to the adapter, the products were then purified and amplified by PCR. The PCR products were heat denatured and circularized (ssDNA circle). DNA nanoballs (DNBs) were generated with the ssDNA circle by rolling circle replication (RCR) to enlarge the fluorescent signals at the sequencing process. The DNBs were loaded into the patterned nanoarrays and single-end read of 50 bp were read through on the BGISEQ-500 platform.