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The conversion coefficient C between relaxation time T2value and pore size was calibrated by a high-pressure mercury injection experiment, low-temperature nitrogen adsorption experiment, and NMR experiment for clarifying the effect of CO2huff and puff on the production characteristics of crude oil in shale micronano pores. Then, the effects of huff and puff rounds, the soaking time, and injection pressure on the seepage law of crude oil in shale pores from the microscopic scale were studied. The results showed that the pore size of the main occurrence space of shale oil was mainly distributed in the range of 1–100 nm. The oil production of matrix/matrix-fractured shale mainly depended on the first two rounds. Combined with MRI imaging technology, in the initial stage, CO2mainly used the crude oil in the surrounding area of the core for matrix shale, and CO2mainly used the crude oil in the fracture and the surrounding area of the fracture for matrix-fractured shale. With the increase of huff and puff rounds, the crude oil inside the core was gradually utilized through CO2diffusion for matrix shale and matrix-fractured shale. Prolonging the soaking time could significantly improve the recovery degree of crude oil in small pores, which increased by 4.21%, but the lower limit of pore size was reduced by only 2–3.4 nm. Increasing the injection pressure could not only reduce the lower limit of pore size, which was reduced by 2.4–3.9 nm, but also effectively improve the recovery degree of crude oil in different pores. Compared with the matrix shale core, the matrix-fractured shale core was more sensitive to the soaking time and injection pressure. The research results could provide some reference and guidance for the field test of CO2huff and puff in shale reservoirs.