GSM8895172: Con3; Sus scrofa domesticus; RNA-Seq
After retrograde perfusion, tissue specimen from the donor's right lung was collected as control group, and the allograft after 8-hour reperfusion was harvested as IRI group. 3 pairs of lung specimen in each group were used for single cell RNA sequencing. Lungs tissues were immediately placed in MACS Tissue Storage Solution (Miltenyi Inc, 130-100-008), and then digested using a lung dissociation kit (Miltenyi Inc, 130-095-927) following the manufacturer's instructions. Cell solution was filtered with 100 μm and 40 μm cell strainers respectively, and underwent RBC lysis using erythrocytes lysate (Biosharp, BL503B). For donor samples, the whole cells were then processed for single-cell RNA sequencing (scRNA-seq). For recipient samples, we performed fluorescence-activated cell sorting in order to obtain more cell types. Cell suspension from transplanted lungs were stained with FITC CD45 antibody (ThermoFisher, 11-9459-42) and Zombie Aqua™ Fixable Viability Kit (Biolegend, 423101). CD45 positive cells were sorted from 2 samples and CD45 negative cells were sorted from 1 sample using a BD FACSAriaTM II flow cytometer (BD Biosciences, USA). scRNA-seq was conducted by BGI Genomics (Beijing, China). Briefly, scRNA-seq library was generated using the DNBelab C Series HT Single-Cell RNA Library kit (MGI, 940-000047-00, China) following the manufacturer's protocol, followed by sequencing on the DNBSEQ-T7 platform at the China National Gene Bank (CNGB).