Inducing metamorphosis

Materials

  • 3~4 days old planula larvae
  • GLWamideII (GNPPGLW-NH2) 1 mg/ml
  • Normal (1 x 3 inch) or wide (2 x 3 inch) glass slides, properly marked with diamond pen (not a marker pen).
  • 10 cm petri dish
  • 1 ml syringe with 25G needle (to smash artemia)

Procedure

  1. Add x1/1000 GLWamide II (final 1 ng/┬Ál) in MFSW (prepare freshly).
  2. Prepare stable and non-disturbing bench/desk. Use a black plastic bag if the surface is not a dark color. It facilitates to see planulae.
  3. Put a glass slide into a plastic dish and place on the stable place. (Keep separate the slide from the wall of the dish.)
  4. Quietly pour 2~3 ml of GLWamideII-MFSW for a normal slide (4~5 ml for double-width slide) using a transfer pipette.
  5. Collect planulae with mouth pipette and put on the glass slides. Planulae will stop swimming immediately (5~10 seconds) and will undergo metamorphosis.
    Inducing_metamorphosis-002-copy.JPG
  6. Leave over a night.
  7. Next morning, transfer slides to a culture tank. Primry polyp is safe out of sea water for short time (5~10 seconds) for transferring from the petri dish to the tank.
    Slides_(primary_polyps)-002 copy.JPG
  8. Some primary polyps start to eat 1~2 days after metamorphosis. Feed smashed artemia, passed through the 25G needle 2~3 times, to primary polyps for several days. Normal feeding will be OK once they form bigger polyps.

Important note: Primary polyp is smaller than normal polyp. They often degrade the polyp body after eating for up to a few times. This is because the primary polyp cannot simply change its body size and a colony makes bigger polyp through degrade-reform process. Reforming new (and bigger) polyp however fails quite often. Only small population of primary polyps, for example 10%, will grow to colonies more than 2 polyps.