title: GSM8264386: LATS1/2 KO replicate 2; Mus musculus; RNA-Seq
library_construction_protocol: The third, fourth, and fifth pairs of mammary glands from each mouse were excised and lymph nodes removed from the fourth pair. Following this, each sample was chopped using a McIlwain tissue chopper. Samples were then dissociated using a solution of 2 mg/mL collagenase A, 0.5 units/mL dispase, 0.01 mg/mL DNAse, and 1x penicillin-streptomycin in DMEM with rotating at 37°C for 1 hour. ACK Lysing Buffer was then used to lyse red blood cells, TrypLE Express with 0.01 mg/mL DNAse was used to dissociate samples to single cells, and samples were strained using a 40 μM cell strainer. Samples were then incubated in 1 μM Calcein Blue, AM for 15 minutes, washed with PBS, and strained into tubes for flow cytometry. Live cells were sorted into a solution of 0.04% BSA in PBS to proceed with processing for single-cell sequencing. Library construction was performed using 10x Genomics protocols (3'v3), including fragmentation, A-tailing, adaptor ligation, and PCR. The quality of the resulting libraries was checked using the Bioanalyzer High Sensitivity DNA Assay (Agilent Technologies).