GSM9177230: Aged; Mus musculus; RNA-Seq
The mice were divided into four groups: The BMF group (experimental group) n=4, the whole-body irradiation (TBI) group (control group) n=3, the normal young mouse group (WT), and the normal aged mouse group (Aged). Specifically, first-generation hybrid B6D2F1 mice were bred by crossing C57BL/6 and DBA/2 mice. These male mice, aged 8-12 weeks, were used as test subjects for the BMF mouse model. Male C57BL/6 mice, aged 8-12 weeks, were used as lymphocyte donors. Male B6D2F1 mice, aged 8 weeks, served as the normal young mouse group, while male B6D2F1 mice, aged 104 weeks, served as the normal aged mouse group. All mice were provided by Beijing Weitong Lihua Experimental Animal Technology Co., Ltd. and were housed in a standard highly hygienic environment. The study was approved by the Experimental Animal Welfare Ethics Committee of Xuanwu Hospital, Capital Medical University (approval number: XW-20210531-1). Cells were first stained with two fluorescent dyes, Calcein AM and Draq7, to precisely determine cell concentration and viability using the BD Rhapsody™ Scanner (BD Biosciences) before antibody-based labeling. Each sample was stained for 30 minutes on ice with 25 AbSeq antibodies targeting major mouse immune markers from the BD Abseq immune discovery panel (Cat. No. 940320, 940107, 940110, 940119, 940132, 940130, 940127, 940168, 940109, 940163, 940131, 940108, 940345, 940137, 940417, 940116, 940496, 940329, 940357, 940437, 940128, 940134, 940165, 940209, 940151). Afterward, the cells were labeled with the respective AbSeq antibodies, washed three times with Stain Buffer (FBS) (BD Biosciences, Cat. No. 554656), and loaded into a BD Rhapsody micro-well cartridge for single-cell capture.Excessive cell capture beads were added to ensure nearly every micro-well contained a bead, and any excess beads were washed away. After cell lysis with lysis buffer, cell capture beads were retrieved, washed, and reverse transcription was performed. The microbeads captured single-cell transcriptomes, along with Ab-tagged index sequences targeting 25 immune cell-associated surface antigens, generating separate cDNA libraries for transcriptome and AbSeq, each containing cell labels and unique molecular identifier (UMI) information. The entire process followed the manufacturer's protocols using the BD Rhapsody cDNA Kit (BD Biosciences, Cat. No. 633773) and the BD Rhapsody WTA Amplification Kit (BD Biosciences, Cat No. 633802). The libraries were sequenced using the PE150 mode (Pair-End for 150 bp read) on the NovaSeq 6000 platform (Illumina).