GSM4633769: E11.5 left lobe replicate 2; Mus musculus; RNA-Seq
Lungs were dissected from CD1 mouse embryos collected at E11.5 in cold PBS. Isolated left lobes were grouped by stage, placed in dispase, and mechanically dissociated with tungsten needles. After 10 minutes in dispase at room temperature, DMEM without HEPES and supplemented with 5% fetal bovine serum (FBS, Atlanta Biologicals) was added, and cell suspensions were passed through a filter with 40-µm-diameter pores. The resultant cell suspensions were then processed by the Princeton Genomics Core Facility. The two sample groups were processed separately. Cells were loaded and processed using the Chromium Single Cell 3' Library and Gel Bead Kit v2 on the Chromium Controller (10X Genomics) following manufacturer protocols. Individual cells were encapsulated in droplets with single gel beads carrying unique barcoded primers and then lysed. Then cDNA fragments were synthesized, barcoded, and amplified by PCR. Illumina sequencing libraries were prepared from the amplified cDNA from each sample group using the Nextra DNA library prep kit (Illumina). The v2 libraries were sequenced on Illumina HiSeq 2500 Rapid flowcells (Illumina) as paired-end 26 + 50 nucleotide reads following manufacturer protocols.