In order to establish an in vitro culture model for male self-compatiblity and -incompatibility in Citrus, we assessed the culture environmental condition and optimal constitution of the liquid culture medium using mature pollen of Hyuganatsu (Citrus tamurana hort. ex Tanaka). A basal medium (Hirano・Hoshino, 2009) was used for examination of the optimum constitution. Mature pollen was cultured in a modified medium containing 0.04% (w/v) CaCl 2 , 0.01% (w/v) H 3 BO 3 , 0.0007% (w/v) KH 2 PO 4 , 0.02% (w/v) yeast extract and 10% (w/v) sucrose with the pH adjusted to 7.0, at a density of 1 × 10 5 pollen grains・mL −1 . These were kept at 25°C under dark conditions. In particular, the addition of CaCl 2 , yeast extract and sucrose to the culture medium was essential for an efficient liquid culture system of mature pollen. In this culture, the nomal germination of pollen grains occurred after 4 hours, and the pollen tubes extended approximately 20 µm per hour. The nomal germination rate of mature pollen was highest (73.6%) after 6 hours of culture. Good growth of pollen tubes was maintained, and they reached 200 µm in length after 8 hours of culture.
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