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Ann Bot. 2006 Jun;97(6):1139-44. Epub 2006 Apr 18.

Isolation of individual egg cells and zygotes in Alstroemeria followed by manual selection with a microcapillary-connected micropump.

Author information

1
Field Science Center for Northern Biosphere, Hokkaido University, Kita 11, Nishi 10, Sapporo 060-0811, Japan. hoshino@exfarm.agr.hokudai.ac.jp

Abstract

AIMS:

To develop a procedure for isolating living egg cells and zygotes from Alstroemeria ovules.

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 24 h 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.

CONCLUSIONS:

The procedures for isolating egg cells and zygotes in Alstroemeria were established, and each egg cell and zygote was captured with a microcapillary.

PMID:
16621859
PMCID:
PMC2803394
DOI:
10.1093/aob/mcl072
[Indexed for MEDLINE]
Free PMC Article
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