GSM5709266: SARSCoV2_Ext_5P_Inf_GEX; Chlorocebus aethiops; RNA-Seq
Cultures were washed with calcium/magnesium-PBS and disassociated with TrypLE (Gibco # 12605010) for 5 minutes at 37°C. Cells were then centrifuged (200 x g for 5 minutes), resuspended in calcium/magnesium free PBS supplemented with 1% BSA and counted. Cells were then filtered through a 40μm FlowMi strainer (ScienceWare # H13680-0040) and counted again. Mock and infected samples were loaded on separate lanes of a 10X Genomics Chromium Controller, according to manufacturer's protocol, for either NextGem Single Cell 3' v3.1 (10X 3') or NextGEM Single Cell V(D)J v1.1 (10X 5'). Single cell RNA sequencing of mock and SARS-CoV-2 infected Vero E6 cells performed with either 10X Genomics Chromium Next GEM Single Cell 3' (10X 3') or 10X Genomics Chromium Next GEM Single Cell V(D)J (10X 5'). Final 10X 3' mock and infected gene expression libraries and 10X 5' infected gene expression libraries were PCR amplified for 16 cycles, while the 10X 5' mock gene expression library was amplified for 14 cycles. 10X 3' libraries were pooled and sequenced according with the following read lengths: read1 28 nt, i7 index 8 nt, read2 130 nt. 10X 5' libraries were also pooled and sequenced with the following read lengths: read 1 26 nt, i7 index 8nt, read 2 132 nt. 10X 5' were also sequenced with our custom extended R1 protocol (read 1 158 nt, i7 index 8 nt, no read 2).