Potato Germplasm Enhancement Laboratory

Obihiro Univerisity of Agriculture and Veterinary Medicine
West 2-11, Inada, Obihiro, Hokkaido 080-8555, Japan

Pollen fertility test by germination ability

Culture medium (for 50 samples)
  5 g Sucrose (20 % sucrose) C12H22O11
  0.25 g Bacto Agar
  2.5 ml 10 x stock solution [0.3 g Ca(NO3)2·4H2O and 0.05 g H3BO3 (50 ppm) in 100 ml distilled water]

  1. Place components of culture medium in a beaker and fill up to 25 ml with distilled water.

  2. Warm to dissolve in a microwave oven.

  3. Pour the dissolved solution on a slide glass to an approximate size of a cover slip using a Pasteur pipette.

  4. Solidify the culture medium in a moisture box (a sealed plastic container with a sheet of wet paper towel on the bottom) at least for 2 hours.

  5. Strew pollen grains on the medium.

  6. Incubate pollen in a moisture box at room temperature for 3-16 hours.

  7. Cut off the edge of the culture medium to the size of 1 cm square.

  8. Place a drop of 1% acetocarmine and cover with a cover slip.

  9. Place the slide glass on a heater, and melt slowly the culture medium (don't boil up).

  10. Remove the slide glass from the heater just after the culture medium is melted.

  11. Observe pollen tube growth under a microscope.