GSM9370542: M48-2; Homo sapiens; RNA-Seq
At the designated time point, media were aspirated, and collagen hydrogels were gently removed from their well and transferred to a 1.7ml tube. Using small surgical scissors, gels were cut into pieces before adding sterilized digestion buffer (2mg/ml Type 7 Collagenase (Worthington) in HBSS) at a 2:1 buffer to gel ratio. Samples were incubated at 37°C for 10 minutes before lightly vortexing and pipetting the samples. After an additional 5-10 minutes, when collagen was no longer visible, cells were pelleted in a microcentrifuge at 300g. Samples isolated directly from droplets were washed off of the plate and centrifuged at 300g. From there all samples were washed with PBS and spun back down before RNA isolation. RNA samples were extracted using RNeasy Mini Kit (Qiagen, 74104). RNA-seq libraries were prepared using standard Poly-A Library Prep (Illumina) and subjected to 151bp paired-end sequencing according to manufacturer's protocols (Illumina NovaSeqXPlus).