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Comparative Detection of Calcium Fluctuations in Single Female Sex Cells of Tobacco to Distinguish Calcium Signals Triggered by in vitro Fertilization
Author(s) -
Peng XiongBo,
Sun MengXiang,
Yang HongYuan
Publication year - 2009
Publication title -
journal of integrative plant biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.734
H-Index - 83
eISSN - 1744-7909
pISSN - 1672-9072
DOI - 10.1111/j.1744-7909.2009.00857.x
Subject(s) - calcium , in vitro , in vitro fertilisation , human fertilization , biology , andrology , microbiology and biotechnology , medicine , genetics , embryo
Double fertilization is a key process of sexual reproduction in higher plants. The role of calcium in the activation of female sex cells through fertilization has recently received a great deal of attention. The establishment of a Ca 2+ ‐imaging technique for living, single, female sex cells is a difficult but necessary prerequisite for evaluating the role of Ca 2+ in the transduction of external stimuli, including the fusion with the sperm cell, to internal cellular processes. The present study describes the use of Fluo‐3 for reporting the Ca 2+ signal in isolated, single, female sex cells, egg cells and central cells, of tobacco plants. A suitable loading protocol was optimized by loading the cells at pH 5.6 with 2 μM Fluo‐3 for 30 min at 30 °C. Under these conditions, several key factors related to in vitro fertilization were also investigated in order to test their possible effects on the [Ca 2+ ] cyt of the female sex cells. The results indicated that the bovine serum albumin‐fusion system was superior to the polyethlene glycol‐fusion system for detecting calcium fluctuations in female sex cells during fertilization. The central cell was fertilized with the sperm cell in bovine serum albumin; however, no evident calcium dynamic was detected, implying that a transient calcium rise might be a specific signal for egg cell fertilization.