The invention relates to the technical field of plant cultivation, in particular to a cunninghamia lanceolata tissue culture seedling transplanting matrix and a preparation method thereof. The transplanting matrix comprises the following components in parts by volume: 1-3 parts of fermented bark, 2-3 parts of fermented wood bran, 4-5 parts of foamed coconut bran and 0.5-1 part of yellow soil. According to the invention, by taking the bark, the wood bran and the coconut bran as main materials, and the yellow soil as an auxiliary material, the cunninghamia lanceolata tissue culture seedling transplanting matrix is prepared, so that the content of the organic matters in the bark is high, the antibacterial property and the stress resistance are achieved, and the content of organic matters in the bark is high, including cellulose, hemicellulose, lignin, or other microelement; the carbon-nitrogen ratio of the wood bran is high, and the air permeability and the water-retaining property are good, the coconut bran is loose in texture, natural decomposition is slow, and the service life of the matrix can be prolonged, the texture of the yellow soil is compact, the loss of moisture and nutrients can be effectively prevented, and the prepared matrix is very suitable for transplanting cunninghamia lanceolata tissue culture seedlings.