The invention provides a method for preparing a cell model capable of detecting mesenchymal stem cell bone differentiation in real time. The method comprises the following steps: S1, mixing sgRNA andCas9 proteins directed at knock-in sites of bone differentiation related genes to form an RNP (ribonucleoprotein) complex; S2, forming to-be-transfected AVV (adeno-associated virus) particles; S3, mixing suspension of stem cells and suspension of the RNP complex and performing electrorotation to obtain a culture subjected to electrorotation; S4, adding the suspension of the to-be-transfected AVV particles into a material subjected to electrorotation and performing transfection to obtain a transfected culture; and S5, extremely diluting the transfected culture and performing monoclonal culture;and performing polymerase chain reaction (PCR) and sequencing, and screening out monoclonal cell strains in which genes are directionally knocked in. Through the technical scheme, according to the method disclosed by the invention, an osteogenic differentiation state of continuous passage cells is monitored in real time in a preparation process of stem cell drugs.