摘要Grape embryo rescue technology is currently the primary method for breeding new seedless grape cultivars.The timing of berry sampling directly impacts the efficacy of this technique.Therefore,achieving efficient,accurate,and non-destructive determination of the optimal sampling time for seedless grape embryo rescue breeding has long been a challenge.This study collected near-infrared spectral data and data on 19 physiological indicators from 2940 grape berries of six grape cultivars at six sampling times to construct a baseline dataset.Remarkably,it was discovered for the first time that pericarp puncture hardness(PPH)is closely associated with the embryo development rate of seedless grape.Subsequently,the optimal sampling times for'Flame Seedless','Ruby Seed-less',and'Jingzaojing'were determined when their PPH reached 720±20 g,990±20 g and 633±20 g,respectively.Then,a total of 840 models for PPH recognition were established and assessed based on their co-efficient of determination(R2)and root mean square error(RMSE).The optimal recognition models for three seedless grape cultivars suitable for embryo rescue—'Flame Seedless','Ruby Seedless',and'Jingzaojing'-were identified as follows:D1+PLSR(R2=0.94,RMSE=42.26),D1+MLR(R2=0.79,RMSE=66.31)and D1+PLSR(R2=0.93,RMSE=47.9).Utilizing the established D1+PLSR or D1+MLR models for PPH,a non-destructive and precise method for sampling seedless grapes during embryo rescue was introduced for the first time.This approach led to a notable increase in the embryo development rate by 15％and enhanced the plantlet rate by 14％.Overall,our proposed strategy provides new perspectives for accelerating the breeding process of new seedless grape cultivars.