Isolation of individual egg cells and zygotes in Alstroemeria followed by manual selection with a microcapillary-connected micropump
Isolation of individual egg cells and zygotes in Alstroemeria followed by manual selection with a microcapillary-connected micropump
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DOI：
10.1093/aob/mcl072
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发表时间：
2006-06-01
期刊：
影响因子：
4.2
通讯作者：
Shinoda, Koichi
中科院分区：
文献类型：
--
作者：
Hoshino, Yoichiro;Murata, Naho;Shinoda, Koichi
center.目的开发一种从Alstroemeria中分离活卵细胞和受精卵的方法尝试从Alstroemeria aurea的胚珠中分离卵细胞和受精卵。ovules.center用透明法对胚珠进行组织学观察，揭示了胚囊和胚珠内卵器的位置和大小。为了分离卵细胞，用手术刀片将胚珠切成切片并用酶溶液处理。随后，在倒置显微镜下使用玻璃针解剖这些胚珠切片。使用连接到微泵的微毛细管收集通过该程序成功分离的卵细胞。对于合子分离，在自花授粉后24小时从子房切除胚珠。通过用酶溶液处理切除的胚珠，随后用玻璃针解剖它们，成功地从胚珠中分离出合子，并用微毛细管收集。分离的合子与花粉管和助细胞相连。卵细胞和受精卵在分离后存活长达2小时，如通过荧光素二乙酸盐测定的。结论建立了用于分离Alstroemeria中的卵细胞和受精卵的程序，并且每个卵细胞和受精卵用微毛细管捕获。staining.center
center dot Aims To develop a procedure for isolating living egg cells and zygotes from Alstroemeria ovules.center dot Scope An attempt was made to isolate egg cells and zygotes from the ovules of Alstroemeria aurea. The ovules were histologically observed using a clearing procedure which revealed the localization and sizes of the embryo sacs and egg apparatus within the ovules. For the isolation of egg cells, ovules were cut into sections with a surgical blade and treated with an enzyme solution. Subsequently, these ovule sections were dissected using a glass needle under an inverted microscope. Egg cells successfully isolated by this procedure were collected using microcapillaries connected to a micropump. For zygote isolation, ovules were excised from ovaries 24h after self-pollination. By treating excised ovules with an enzyme solution and subsequently dissecting them using a glass needle, zygotes were successfully isolated from the ovules and collected with a microcapillary. The isolated zygotes were associated with pollen tubes and one of the synergids. Egg cells and zygotes were viable for up to 2 h following isolation, as determined by fluorescein diacetate staining.center dot Conclusions The procedures for isolating egg cells and zygotes in Alstroemeria were established, and each egg cell and zygote was captured with a microcapillary.