GSM8571976: Ferret 4626 Saline Replicate 1; Mustela putorius furo; RNA-Seq
Lung nuclei from saline and bleomycin injured ferrets were isolated using a CST buffer as described previously. Snap frozen lung tissues (50 mg) were immediately placed into a nuclei isolation using CST buffer (20 mM Tris-HCl pH 7.5, 292 mM NaCl, 2 mM CaCl2, 42 mM MgCl2, 1% CHAPS, and 2% BSA) and homogenized using a dounce homogenizer to mechanically dissociate nuclei from cells. The resulting homogenate was filtered through a 40 µm strainer, and nuclei were pelleted by centrifugation at 300 × g for 2 minutes at 4°C. Nuclei were resuspended in 0.04% BSA buffer to a final concentration of 1,000 nuclei/µl. Single-nuclei RNA sequencing (snRNA-seq) libraries were prepared using the 10x Genomics 5' Kit v2, following the manufacturer's protocol (10X Genomics). Libraries were sequenced on an Illumina Nextseq 550 platform using 75-cycle paired-end sequencing, using 8bp for index read 28bp for R1 and 56bp for R2.