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用两个解剖针挤压胡萝卜花粉使其破裂释放出精细胞。用酶解-解剖方法分离胡萝卜胚囊中的卵细胞、助细胞和中央细胞。胡萝卜胚珠先在酶液中酶解40~50min,然后将其转移到不含酶的分离液中用解剖针解剖胚珠。将胚珠的合点端切破,轻轻挤压胚珠的珠孔,卵细胞、助细胞和中央细胞即可逸出。在最佳条件下,20min可从20个胚珠中分离出5个卵细胞。对分离胚囊细胞的渗透压和酶液成分进行了筛选。分离出的卵细胞用显微操作仪收集。胡萝卜精、卵细胞的成功分离为在双子叶植物中进行离体受精探索创造了条件。
Squeezing carrot pollen with two dissecting needles ruptures to release sperm cells. Isolation of egg, accessory and central cells from carrot embryo sacs by enzymatic and dissection methods. The carrot ovule is first enzymatically digested in the enzyme solution for 40-50 min and then transferred to an enzyme-free separation solution to dissect the ovule with a dissection needle. The oval end of the joint cut, gently squeeze the ovule beads, eggs, cells and central cells can escape. Under the optimal conditions, 5 egg cells were isolated from 20 ovules in 20 min. The osmotic pressure and enzyme composition of the isolated embryo sac cells were screened. The isolated egg cells were collected using a micromanipulator. The successful isolation of carrot sperm, egg cells creates the conditions for ex vivo fertilization exploration in dicotyledons.