Clearing Arabidopsis ovule materials with the Hoyer's solution

Ref: Liu C.M. and Meinke D.W. (1998) The titan mutants of Arabidopsis are disrupted in mitosis and cell cycle control during seed development. Plant J. 16, 21-31

This protocol can be used for clearing intact seeds of arabidopsis, which can then be observed under Normarski optics. This is an efficient way to analyse embryo and endosperm development without sectioning

  1. To prepare the modified Hoyer's solution:
    bullet7.5g gum arabic (Sigma, G9752)
    bullet100g chloral hydrate (Sigma, C8383, controlled substance)
    bullet5ml glycerol (Sigma, G6279)
    bullet60ml water
    bulletdissolve by stirring for 3-5 hrs, or overnight.
  2. Open the siliques with a pair of forceps and a sharp needle, pick up the intact ovules with the needle.
  3. Put 2-3 drop of Hoyer's on a slide.
  4. Drop the ovules into the Hoyer's. 20-30 ovules for each sample.
  5. Gently cover the sample with a cover slip.
  6. Keep the slides at room temperature for 15' to overnight depending on the developmental stages of the ovules.
  7. Observe the samples with a microscope equiped with Nomarski optics.