Illumina NovaSeq 6000 sequencing; A high-resolution single-cell multiomic atlas of the human fetal lung - Organoid
A high-resolution single-cell multiomic atlas of the human fetal lung - Organoid
Protocol carried out by HDBR to extract specimen from a fetal donor.  Human fetal lung organoids were derived and maintained as previously described (Nikolic et al., 2017). In brief, human foetal lung tissues were treated with Dispase (8 U/ml Thermo Fisher Scientific, 17105041) at room temperature (RT) for 2 min to digest mesenchymal connections. Mesothelium and mesenchymal cells were carefully removed by needles. Branching epithelial tips were micro-dissected by needles, transferred into Matrigel (356231, Corning) and seeded in a 24 well low-attachment plate (M9312-100EA, Greiner) with 4-5 tips per 50 l Matrigel dome per well. The plate was incubated at 37C for 5-10 min until the Matrigel domes solidified. 600 l of self-renewing differentiation medium was added. Organoids were cultured under standard tissue culture conditions (37C, 5% CO2), maintained in self-renewing medium and passaged by mechanically breaking using P200 pipettes every 4-7 days. Organoids were then fully dissociated into single cells by incubation with prewarmed TrypLE (Thermo Fisher Scientific, 12605028) for 15-20 min with trituration every 5 min.  Libraries were prepared according to the standard protocol of the Chromium 5'V1 kit.