The invention provides a preparation method of high-purity paclitaxel. The preparation method comprises the following steps: (1) establishing a stable taxus chinensis forming layer stem cell culture system; (2) adding taxus chinensis endophytic fungi extract liquid into a suspension culture medium, inoculating taxus chinensis cells, performing the ventilation culture for 6 to 8 days under an artificial lighting condition, and adding jasmonic acid methyl ester and a clinacanthus nutans extract into the liquid suspension culture medium to continue the culture; (3) collecting the taxus chinensiscells, and performing the coarse extraction of paclitaxel; (4) purifying a crude extract; and (5) enabling a concentrated solution II to pass through a C18 high-performance liquid phase chromatographic purification column, eluting by using methanol, collecting and combining paclitaxel elution parts, decompression evaporating the solvent methanol, crystallizing and releasing the paclitaxel, centrifugally collecting crystals, collecting and combining the paclitaxel elution part, decompression evaporating the solvent methanol to crystallize and precipitate the paclitaxel, centrifugally collectingthe crystals, and vacuum freeze drying the crystals, thus obtaining the high-purity paclitaxel concentrated solution II. The preparation method has the characteristics of high induction success rate,high paclitaxel extraction purity and the like.