title: GSM7136514: Replicate 1, GFP low; Mus musculus; RNA-Seq
library_construction_protocol: Tumours were dissociated using papain and cells sorted based on tdTomato and GFP fluorescence into 0.04% BSA in PBS. Cells were processed according to 10X Genomics User Guide (Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (Dual Index)). Cells were resuspended with RT Master Mix and loaded onto Chip G along with the barcoded gel beads and partitioning oil in order to generate the gel bead-in-emulsion (GEM) on the Chromium Controller. Once complete, the GEMS are loaded onto a thermocycler to complete the RT inside each GEM to produce cDNA, which contain Illumina R1 primer sequence primer sequence, Unique Molecular Identifier (UMI) and the 10X Barcode. Silane Dynabeads are used to clean up the pooled barcoded cDNA. The cDNA is then amplified and a quick QC is conducted to look at the fragment sizes. 10ul of the amplified cDNA is taken forward for the library construction which involves 3 steps of fragmentation (to obtain fragments around 450bp), ligation (to add the Illumina p5 and p7 primers) and Indexing (to add unique sample index to each sample). These steps are always followed by a SPRI Select bead purification. Once Indexing is complete the libraries were pooled and submitted for sequencing.