GSM8999721: d9435_d7_mal; Homo sapiens; RNA-Seq
For bulk RNA-seq, total RNA was extracted from organoid lysates using the RNeasy Mini Kit (QIAGEN) following the manufacturer's instructions. Samples were quantified with a NanoDrop (Thermo) before storing at -80°C. For single cell RNA sequencing, Mock, ZIKVPR, ZIKVMAL, and ZIKVUG infected organoids at 7 dpi were manually dissociated into a single cell suspension using the Embryoid Body Dissociation Kit (Miltenyi Biotec). The manufacturer's protocol was followed with these adjustments: 1 ml of enzyme mix was used per 3 organoids, 1 ml of ice-cold Maturation media was used to wash the pre-separation filter, and the cell suspension was centrifuged at 300×g for 10 min to better pellet neurons. scRNA-seq was performed using the Chromium Next GEM Single Cell 3' Kit v3.1 (10x Genomics) following the manufacturer's protocol. Single cell suspensions were loaded onto the Chromium Controller (10x Genomics) for droplet formation. scRNA-seq libraries were prepared using the Chromium Single Cell 3' Reagent Kit (10x Genomics). Samples were sequenced by the MIT Genomics and Informatics Facilities core.