Abstract: The effect of temperature and food concentration on resting egg formation of freshwater rotifer Brachionus calyciflorus was studied by accumulative population culture under 9 treatments of 3 temperature gradients (20℃, 25℃ and 30℃) and 3 food concentration gradients (1.5 ×106cells / mL,3.0 ×106cells / mL and 6.0 ×106cells /mL). The results showed that under the culture condition of temperature 20℃ and food concentration 6.0×106cells / mL, the production and formation efficiency of resting egg were 2032.33±141.50 ind./(10mL. 13d) and 2.61±0.18 ind./106cells algae, respectively, and were both biggest among all the conditions. This condition was optimal for mass production of resting eggs. At temperature 25℃ and 30℃, the food concentration of 6.0 ×106cells / mL and 3.0 ×106cells / mL was also optimal for the production of resting eggs, respectively. At the temperature 20℃, the mixis rate was bigger than that of 25℃ and 30℃. There was no significant effect of temperature,food concentration and their interaction on the fertilization rate.