Lin−PU.1dimGATA-1− (LPG) defines hematopoietic stem cells with long-term multilineage reconstitution activity

We isolated single cells from Spi1-eGFP&Gata1-mCherry double knock-in mice. Briefly, the bone marrow tissues were flushed from adult mouse femurs with PBS buffer and passed through 40 μm strainers, followed by flow cytometer (BD FACS Aria II), then LSK cells and Lin- PU.1dim GATA1- cells were collected and counted. With help from CapitalBio Technology Inc. (Beijing, China), single-cell RNA-seq libraries were constructed according to the instructions provided by 10x Genomics that accompany single cell 3′ Library and Gel Bead Kit V3 (10x Genomics, 1000075). The cDNA library was then generated, amplified, and assessed for quality control using the Agilent 4200, and Single-cell RNA sequencing was further performed on the Illumina Novaseq6000 sequencer. Read pre-processing was performed using the 10 × Genomics workflow.