The effects of planting pattern of parental lines and the pollination mode of honey bees on the fruit development and seed yield of hot pepper (Capsicum annuum L.) grown in the net-house were studied in order to understand the potential of using bee pollination in seed production. We found that cytoplasmic male sterile line C was suitable for honey bee pollination in seed production due to its larger flowers, more nectar secretion, and higher honey bee foraging frequency compared to other lines tested. When relying on honey bees to do pollination in the closed net-house, the male and female parents grown in 1:1 ratio within the same row could obtain more seed production. Both fresh pollens and liquid nitrogen preserved pollens could be used for honey bees to carry them for pollination and make successful fruit-sets and seed production, but using bee pollination this way yielded less seeds than using hand-pollination.