title: GSM7785049: Mock_0hpi_mCD300lf-hCaco2, scRNAseq; Homo sapiens; RNA-Seq
library_construction_protocol: MNV was infected at MOI 5 titrated by RAW264.7 cells for 1 hour in a CO2 incubator at 37 C°. The UV-inactivated MNV was also inoculated on the cells for negative control on the infection. For mock controls, the MEM supplemented with GlutaMax and 20% FBS without the virus were added on the cells. The inoculums were removed from the infected cells and they were washed with PBS(-). The media were added on the infected cells and they were incubated with a CO2 incubator at 37 C° for 3 hour, 7 hours or 11 hours. The cells were detached by 0.25% Trypsin-EDTA and centrifugated with 250xg for 5 min. The cell pellets were resuspended with 1.5 mL of the media and the cells were passed through Flowmi cell strainers. The filtered cells were quantified by Acridine Orange/Propidium Iodide Stain and then the Luna fx7 cell counter. The cells were further washed with PBS(-) including 0.04% BSA for 2 times after centrifugation with 150xg for 3 min. The cell pellets were resuspended with 500uL of the PBS(-) including 0.04% BSA and quantified by the Luna fx7 cell counter again. The cells were diluted to 1,000 cells per microliter with the PBS(-) including 0.04% BSA. Library was performed according to the manufacter's instructions (single cell 3' v3.1 protocol, 10x Genomics). Briefly, mCD300lf-hCaco2 cells were resuspended in the master mix and loaded with partitioning oil and gel beads into the chip to generate the gel bead-in-emulsion (GEM). The polyA RNA from the cell lysate contained in single GEMs was retrotranscripted to cDNA, which contains an Ilumina R1 primer sequence, Unique Molecular Identifier (UMI) and the 10x Barcode. The pooled barcoded cDNA was then cleaned up with Silane DynaBeads, amplified by PCR and the appropriate sized fragments were selected with SPRIselect reagent for subsequent library construction. During the library construction Ilumina R2 primer sequence, paired-end constructs with P5 and P7 sequences and a sample index were added.