Objective：CRISPR/Cas9 gene editing technology was employed to knockout the PIN1 gene of adult neural stem cells（NSCs）and finally established a PIN1 gene knockout adult neural stem cell line. Methods：The tissue from the subventricular zone of 8-week-old C57BL/6 mice was dissected and cultured in vitro；single guide RNA（sgRNA）targeting PIN1 gene in mice was designed，and the PX330 plasmid was used as the skeleton to construct PIN1 Cas9 target vector. PIN-knockout monoclonal cells were obtained after transfection and puro screening. The expression level of PIN1 protein was verified by immunofluorescence and Western blot. The expression of nestin was used to identify the neural stem cell characteristics. The differentiation ability to neuronal lineage was determined by beta Ⅲ tubulin immunofluorescence. Results：Cas9/sgRNA expression vector of PIN1 gene was successfully constructed，and nine PIN1 knockout neural stem cells were cloned after transfection. Immunofluorescence and Western blot showed no expression of PIN1 protein，immunofluorescence showed positive expression of nestin in PIN1 knockout neural stem cells，and neural stem cells could differentiate into beta Ⅲ tubulin positive neurons in differentiation culture. Conclusion：CRISPR/Cas9 gene editing technology can realize the editing of PIN1 gene in mouse adult neural stem cells. After knock-out of PIN1 gene，PIN1 protein is not expressed，but neural stem cells still partially express the specific marker nestin and have the ability to differentiate toward neurons.