The heterogeneity of human hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) under stress conditions such as ex vivo expansion is poorly understood. Here, we report that the frequencies of SCID-repopulating cells were greatly decreased in cord blood (CB) [CD34.sup.+] HSCs and HPCs upon ex vivo culturing. Transcriptomic analysis and metabolic profiling demonstrated that mitochondrial oxidative stress of human CB HSCs and HPCs notably increased, along with loss of stemness. Limiting dilution analysis revealed that functional human HSCs were enriched in cell populations with low levels of mitochondrial ROS (mitoROS) during ex vivo culturing. Using single- cell RNA-Seq analysis of the mitoROS low cell population, we demonstrated that functional HSCs were substantially enriched in the adhesion GPCR G1-positive ([ADGRG1.sup.+]) population of [CD34.sup.+][CD133.sup.+] CB cells upon ex vivo expansion stress. Gene set enrichment analysis revealed that HSC signature genes including MSI2 and MLLT3 were enriched in [CD34.sup.+][CD133.sup.+][ADGRG1.sup.+] CB HSCs. Our study reveals that ADGRG1 enriches for functional human HSCs under oxidative stress during ex vivo culturing, which can be a reliable target for drug screening of agonists of HSC expansion.