GSM5857307: Dropseq-T2D4; Homo sapiens; RNA-Seq
We performed Drop-Seq following the protocol as previously described (Macosko et al., 2015). Briefly, three pump-controlled syringes with cell suspension (200,000 cells/mL), barcoded beads in lysis buffer (120,000 beads/mL), and droplet generation oil were connected to a microfluidic device under microscope supervision. During droplets generation, we set the cell and bead flow speed at 4,000uL/hr, and the oil speed at 15,000uL/hr. The droplets are collected into 50mL falcon tubes (usually less than 5mL). Under this setting, most droplets have at most one beads or one cell. Following droplet breakage,we performed 1st strand cDNA synthesis on beads followed by SMART-PCR (Macosko et al., 2015). Finally, the resulting full-length cDNA library are prepared for sequencing exactly following the published Drop-Seq protocol (Macosko et al., 2015).