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Programmed Cell Death Ligand 1 (PD-L1) is a trans-membrane protein that attenuates the host immune response to tumor cells. PD-L1 ligand on the surface of the cancer binds to PD-1 transmembrane receptors on T cells and exhausts T cells function. Using CRISPR-Cas9 genome editing, we generated biallelic PD-L1 mutants in human induced pluripotent stem cells (hiPSCs). The PD-L1 homozygous-knockout hiPSCs retained their normal morphology, gene expression, and in vivo differentiation potential.