In this paper, the sensitivity factors of CO2 huff-n-puff for multifractured horizontal wells (MFHWs) in tight oil reservoirs were investigated through an experimental test and numerical simulation. The pressure-volume-temperature (PVT) experiment and the slim tube experiment are used to understand the interaction mechanism between CO2 and crude oil, and the minimum miscibility pressure (MMP) of the CO2-crude oil system is 17 MPa. The single-well model was firstly established to analyze the sensitivity factors on production performance of MFHWs by using CO2 huff-n-puff. The controlling factors of CO2 huff-n-puff for MFHWs in tight oil reservoirs were divided into three categories (i.e., reservoir parameters, well parameters, and injection-production parameters), and the impact of individual parameter on well performance was discussed in detail. The range of reservoir parameters suitable for CO2 huff-n-puff of MFHWs is obtained. The reservoir permeability is from 0.1 mD to 1 mD, the reservoir thickness changes from 10 m to 30 m, and the reservoir porosity is from 7% to 12%. Based on the reservoir parameters of the target reservoir, the reasonable well and fracture parameters are obtained. The sensitivity intensity was followed by the horizontal well length, fracture conductivity, fracture spacing, and fracture half-length. CO2 injection-production parameters are further optimized, and the sensitivity intensity was followed by the single-cycle cumulative CO2 injection rate, the soaking time, the injection rates, and the production rates. It provides a reference for parameter optimization of CO2 huff-n-puff for MFHWs in tight oil reservoirs.