Normal estrous cycle is crucial for porcine reproduction, and microRNA is closely related to regulation of estrous cycle in porcine ovaries. In this study, we found that the expression of miR-214 in porcine ovaries was higher than in many other tissues, and miR-21 expression in ovaries was significantly higher than in the uterus and pituitary. Meanwhile, miR-21 was upregulated and miR-214 was downregulated in the ovaries of high litter size (YH) pigs compared with low litter size (YL) pigs. Moreover, the lowest expression of miR-21 and miR-214 occurred on Days 14 and 7 of the estrous cycle and was expressed at greater levels in the granulosa cells of subordinate follicles than in dominant follicles on Day 3 of the estrous cycle. Bioinformatics analysis showed that miR-21 and miR-214 might target several genes that involved in the mTOR signaling, apoptosis, and steroid biosynthesis pathways, and they play important roles in maintaining the porcine estrous cycle. The qPCR and western blot analysis indicated that miR-214 inhibited the expression of SCARB1 gene in the transcriptional level, but not affected the SCARB1 gene's protein level. Our research findings indicated that miR-21 and miR-214 played important roles in reproduction regulation during porcine estrous.