Abstract: Using Muscovy as male parent, white Kaiya duck as female parent, the fertilization rate of mule duck was analyzed by different insemination parameters (insemination depth, insemination amount, insemination time and insemination interval) .The results showed that the fertilization rate of 6cm depth insemination was significantly higher than that of 3cm depth insemination.The average fertilization rate of 3cm depth insemination was 71.01% and the average fertilization rate of 6cm depth insemination was 76.43% by using different insemination amount, and there was no significant difference among different insemination amount.The fertilization rates were 71.63% and 70.46%respectively in morning and afternoon, showing no significant difference on different insemination time.The fertilization rates of 2days and 4days insemination interval were significantly higher than the fertilization rate of 6days insemination interval, and there was no significant difference between 2days and 4days insemination interval.The result of this study provides the theoretical basis and reference value for mule duck artificial insemination.