GSM7957490: sample.hoxcep120het_2; Mus musculus; RNA-Seq
For analysis of GFP-positive UB cells with and without centrosomes during embryogenesis, kidneys of wildtype and Hoxb7-Cep120 mice were isolated at E13.5 and dissociated into single cell suspension as previously described (Jain et al., 2014). Briefly, E13.5 embryonic kidneys were dissected and collected in 1.5 ml tube. Collected kidneys were suspended with Trypsin-EDTA solution (Sigma, T3924) with 200 g/ml DNase I (Sigma) and incubated for 10 min at 37°C. After the incubation the specimens were triturated with a P-200 pipette and rinsed once with DMEM/F12 containing 10% FCS to inactivate Trypsin. The cell preparation was then treated with collagenase dissolved in DMEM/F12 (1 mg/ml) and incubated at 37°C for 10 min. The specimens were again triturated and fully dissociated cells were rinsed twice in 500 µl of PBS/ 5% FCS and subjected to Fluorescence active cell sorting (FACS). FACS sorting (Sony Synergy-HAPS 1, 100 micron) was performed using the GFP channel. ds-cDNA was prepared using the SMARTer Ultra Low RNA kit for Illumina Sequencing (Takara-Clontech) per manufacturer's protocol