Chin Bull Bot ›› 2009, Vol. 44 ›› Issue (03): 345-350.doi: 10.3969/j.issn.1674-3466.2009.03.012

• 技术方法 • Previous Articles     Next Articles

Isolation of Egg Cells from Allium fistulosum

Mingxia Jian, Yanan Zhang, Yaying Wang, Huiqiao Tian*   

  1. School of Life Sciences, Xiamen University, Xiamen 361005, China
  • Received:2008-06-17 Revised:2008-10-01 Online:2010-11-03 Published:2009-05-01
  • Contact: Huiqiao Tian

Abstract:

Viable egg cells of Allium fistulosum were isolated by enzymatic digestion and mechanical dissection. The ovules were incu-bated in an enzymatic solution containing 0-0.8% pectolyse Y-23, 0-1.2% pectinase (Serva), 0-1.2% cellulase (Onozuka R-S), 0-1% hemicellulase (Sigma) and 6%-12% mannitol for 20-40 min to remove the outer integument. Then ovules were tr-ansferred into an isolating solution without enzymes for mechanical dissection. The ovules were cut from the middle part, and the micropylar part was gently extruded by use of a dissection needle. In general, the three cells of the egg apparatus spilled out from the cut end of the ovule. The enzymatic solution containing 0.2% pectolyse Y-23, 0.8% pectinase (Serva), 0.8% cellulase (Onozuka RS), 0.5%hemicellulase (Sigma) and 10% mannitol was best for isolating about 18 egg cells from 30 ovules in 2 h. The isolated egg cells could be used for in vitro fertilization to explore the fertilization mechanism and for egg development with molecular biological methods.

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