Choroid plexus organoids as a model of the blood-cerebrospinal fluid barrier to study Streptococcus suis infection
Choroid plexus organoids as a model of the blood-cerebrospinal fluid barrier to study Streptococcus suis infection
The total RNA was isolated from ChP organoids (challenged with S. suis ± Plg or Control) using the RNeasy Mini Kit (Qiagen). Three replicates were used for each condition and each replicate consisted of  three ChP organoids collected into a 15 mL tube. Organoids were dissociated using Accutase (STEMCELL, 07920) by pipetting up and down 10 times at 5 min intervals. Cells suspension was pelleted at 300 × g for 5 min, resuspended in 350 µL of Buffer RLT, and thoroughly disrupted by vortexing. 350 µL of 70% ethanol was added to tube and up to 700 µL of the sample was transferred to an RNeasy Mini spin column placed in a 2 mL collection tube.  Subsequently, genomic DNA was eliminated from RNA samples using DNA-freeTM DNA Removal Kit (Invitrogen AM 1906) following the manufacturer's recommended protocol. Messenger RNA was purified from total RNA using poly-T oligo-attached magnetic beads. First-strand cDNA was synthesized using random hexamer primers, followed by the second-strand cDNA synthesis using dTTP for non-directional library construction