In view of the problems of low matrix permeability and low oil recovery in shale reservoirs, CO2 huff and puff technology is considered as an effective method to develop shale oil reservoirs. However, the production behaviors of actual shale reservoirs cannot be reproduced and the EOR potentials cannot be evaluated directly by scaled models in the laboratory. Conventional CO2 huff and puff has problems such as early gas breakthrough and gas channeling leading to inefficient development. In this article, with the help of a three-dimensional experimental simulation apparatus, a new method of CO2 huff and puff with a horizontal well assisted by pumping production and water injection disturbance is developed. The dynamic characteristic, pressure field distribution of soaking and the enhanced oil recovery effect are comprehensively evaluated. The results show that the soaking stage of CO2 huff and puff can be divided into three stages: differential pressure driving, diffusion driving and dissolution driving. According to the pressure field distribution, after water injection disturbance, the fluctuation boundary and distribution of pressure becomes more stable and uniform and the sweep rate is greatly improved. Water injection disturbance realizes the combination of CO2 injection energy enhancement and water injection energy enhancement and the CO2 injection utilization rate is improved. It has the dual effect of stratum energy increase and economic benefit. The new huff and puff method can increase the oil recovery rate by 7.18% and increase the oil–gas replacement rate to 1.2728, which confirms the potential of horizontal well pumping production and water injection disturbance-assisted CO2 huff and puff technology to improve oil recovery.