Illumina HiSeq 2500 sequencing; Single-cell RNA sequencing of mouse germ cells of the trans-chromosomal mouse model compared to wild type littermates and wild type mice
Single-cell RNA sequencing of mouse germ cells of the trans-chromosomal mouse model compared to wild type littermates and wild type mice
To assay the robustness of the meiotic cell division process, we profiled germ cells of the trans-chromosomal mouse model (Tc1) (containing Tc(HSA21)1TybEmcf genotype) that carries a copy of the human chromosome 21. The Tc0 genotype is the same as Tc1 but did not inherit the human chromosome 21. So these are littermates of Tc1 that were breed with F1 animals (129S8 x C57BL/6J) but lost the chromosome. Spermatogenic cell populations were isolated from adult mouse testes as described in Ernst et al. (2016). In brief, the albuginea was removed and tissue was incubated in dissociation buffer containing 25mg/ml Collagenase A, 25mg/ml Dispase II and 2.5mg/ml DNase I for 30 minutes at 37C. Enzymatic digestion was quenched with Dulbecco's Modified Eagle Medium (DMEM, Gibco) supplemented with 10% Fetal calf serum (FCS, 10270106, Gibco). Mouse testes were enzymatically dissociated and 34 ul of single-cell suspension at a concentration of ~297.000 cells/ml was loaded into one channel of the ChromiumTM Single Cell A Chip (10X Genomics®), aiming for a recovery of 4000-5000 cells. Single-cell barcoding, cDNA synthesis and library preparation were performed following manufacturer's instructions according to Chromium Single Cell 3′ Library & Gel Bead Kit v2, 16 rxns (Product Code: 120237)
Experimental Factor: age
67
Experimental Factor: genotype
wild type genotype
Experimental Factor: strain
C57BL/6J