[Objective]The aim was to determine the collection standard of cotton ovules on different days before flowering,and then to more clearly study the changes in gene expression of cotton ovules before and after blooming.[Method]The standard to distinguish ovules in Xinluzao 13 on different days before flowering was defined by the length and width of alabasterum and sepal.[Result]The alabasterum length increased gradually with approaching of flowering time,while other parameters did not show the same tendency.Therefore,the alabasterum length was chosen as the standard to determine the ovules on different days before flowering.The results of statistics analysis revealed that the alabstrums with length of 1.0-1.2,1.3-1.5 cm and more than 1.7 cm respectively were determined as the standards for ovules collection on 5,3 and 1 d before flowering.[Conclusion]This study will provide not only the reference data for the collection of ovules sample in the following experiment,but also the basis for more precisely detecting the changes in gene expression of ovules before and after flowering.