GSM4743595: Colon organoids at 12hpi Targeted TAP-seq; Homo sapiens; RNA-Seq
2D seeded  organoids harvested after 0 (mock), 12 and 24 hours post-infection were washed in cold PBS and incubated in TrypLE Express for 25 min at 37°C. When microscopic examination revealed that cells had reached a single cell state, they were resuspended in DMEM/F12 and spun at 500 xg for 5 mins. Supernatant was removed and the cell pellet was resuspended in PBS supplemented with 0.04% BSA and passed through a 40-μm cell strainer. For scRNAseq - Single-cell suspensions were loaded onto the 10x Chromium controller using the 10x Genomics Single Cell 3' Library Kit NextGem (10x Genomics) according to the manufacturer's instructions. For TAP-seq, Targeted scRNA-seq was performed as previously described, except for using amplified cDNA from the 10X Genomics 3' scRNA-seq protocol as input material. In short, 10 ng of amplified cDNA were used as input for the outer primer PCR and amplified with 10 PCR cycles. A second seminested PCR using 10 ng of Ampure purified outer PCR as input was performed with inner primer mix and 7 cycles of PCR. Then, a third PCR was done adding Illumina adapters