Palser B F, Rouse J L, Williams E G
Department of Biological Sciences, Rutgers University, Piscataway, New Jersey 08855.
Biotech Histochem. 1992 Jul;67(4):207-18. doi: 10.3109/10520299209110068.
Using ovule clearing, more than 33,600 ovules of Rhododendron nuttallii T. W. Booth (Ericaceae) were examined for megagametophyte and early postfertilization stages at daily intervals from anthesis until 3 weeks after pollination. Pretreatments with amyloglucosidase to digest integumentary and gametophyte starch and Stockwell's bleach to remove tannins from the integumentary epidermis were necessary. Ovules were cleared by a combination or modifications of Herr's four-and-a-half or Stelly's hemalum-methyl salicylate techniques and were observed using differential interference contrast optics. The method proved suitable for large-scale quantitative studies of ovule development and fertilization. A general protocol is suggested as a starting point for ovule clearing studies.
利用胚珠透明法，从花期开始直至授粉后3周，每天对33600多个努氏杜鹃（杜鹃花科）胚珠进行检查，以观察大孢子体和受精后早期阶段。需要用淀粉葡糖苷酶进行预处理以消化珠被和配子体淀粉，并用斯托克韦尔漂白剂去除珠被表皮中的单宁。胚珠通过对赫尔的四点半法或斯特利的苏木精-水杨酸甲酯法进行组合或改进来进行透明处理，并使用微分干涉对比光学显微镜进行观察。该方法被证明适用于胚珠发育和受精的大规模定量研究。建议采用一个通用方案作为胚珠透明研究的起点。
