Wheeler Lab
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    Planaria husbandry

    Created
    Jul 8, 2024 3:41 PM
    Status
    Complete
    Category
    Planaria
    • Changing water
    • Materials
    • Protocol
    • Feeding planaria
    • Materials
    • Protocol
    • Preparing planaria food
    • Materials
    • Protocol

    Changing water

    Every 2 days and after each feeding

    Materials

    • Old Tupperware dish
    • New Tupperware dish
    • Fresh APW (recipe below)
    • 100 mL beaker
    • Small beaker
    • Spray bottle
    • Transfer pipette
    • Gloves

    Artificial Pond Water Recipe (1 L bottle, NIH recipe)

    • 500 µL FeCl3
    • 2,500 µL CaCl2
    • 2,500 µL MgSO4
    • 1,250 µL Phosphate Buffer
    • Fill to 1 L with DI water
    • Label with initials, pH, date, and specify use for planaria only

    0.75X Montjuic salts recipe:

    Salt solution
    Initial concentration
    Final concentration
    Volume added
    NaCl
    5 M
    1.6 mM
    320 µL
    CaCl2
    1 M
    1 mM
    1 mL
    MgSO4
    1 M
    1 mM
    1 mL
    MgCl2
    1 M
    0.1 mM
    100 µL
    KCl
    1 M
    0.1 mM
    100 µL
    NaHCO3
    -
    1.2 mM
    .1008 g
    • Add Milli-Q water to 1.33 L

    Because each 6-8 mm planarian requires 1 mL of water, we can place about (source):

    • 5 worms in Small Petri dish (35 x 10 mm, 5 mL)
    • 10 worms in Medium Petri dish (60 x 15 mm, 11 mL)
    • 50 worms in Big Petri dish (100 x 15 mm, 50 mL)
    • 100 worms in Extra Small Bowl Ziploc® (9 cm diameter x 5 cm depth, 150 mL)
    • 500 worms in Small Square Ziploc® (12 cm x 12 cm x 6 cm, 600 mL)
    • 1000 worms in Large Rectangular Ziploc® (15 cm x 26 cm x 8 cm, 1300 mL)

    Protocol

    1. Pour ~250 mL of fresh APW into the new container.
    2. Pour as much as possible of the old water into the 100 mL.
      1. If there aren’t any planaria that fell in, dump water down the drain.
      2. If there are planaria, use a transfer pipette to move them to the fresh container.
        1. Try not to transfer any food particles/waste when moving the planaria.
    Pouring old water into beaker.
    Pouring old water into beaker.
    Transferring planaria from old water into fresh container using pipette.
    Transferring planaria from old water into fresh container using pipette.
    1. Fill a spray bottle (labelled planaria waste water) with some of the old APW water, and tipping the old container, spray the planaria that stick down to the bottom.
    Filling spray bottle with used water.
    Filling spray bottle with used water.
    Spraying planaria down to bottom of container to empty into the beaker.
    Spraying planaria down to bottom of container to empty into the beaker.
    1. Repeat steps 2 and 3 until there are no planaria left in the old container and all of them are transferred in the new container.
    2. If changing after feeding, remove any large chunks of egg yolk from new container
    3. Label and seal the container with a holed lid cover
    ✅
    This protocol shows cleaning after a feeding (egg leftovers shown) - When cleaning after not feeding, there shouldn’t be any large particles in the old water.
    ⚠️
    We have multiple populations of planaria which have distinct genetic histories. It is very important not to mix these populations. Change gloves and materials in between populations, if necessary.

    Feeding planaria

    Once a week

    Materials

    • Frozen egg yolk
    • Spatula

    Protocol

    1. Remove one frozen cube of egg yolk smoothie (around pea sized amount)
      1. If the cube is broken, scoop enough to fill the tip of the spatula
    2. Place into the middle of the container
    3. If needed, break up chunks with spatula
    image
    1. Allow planaria to eat ~1-2 hrs
    2. After feeding, change the water (Planaria husbandry - Changing water)

    Preparing planaria food

    Complete as needed

    Materials

    • 1000 mL beakers (2)
    • Hot plate
    • Egg (1)
    • Tap water
    • Ice
    • Aluminum foil
    • Paper towel
    • Cutting utensil
    • Dish
    • Spatula
    • Red mold
    • Freezer

    Protocol

    ✅
    Use hot plate in molecular bio suite, 1000 mL beakers found in cabinet above hot plate location. Using more than 1 egg gives plenty of leftovers - freeze any unused product
    1. Fill beaker 3/4 way full of water
    2. Place 1 egg into water
    3. Turn heat on to 10, bring water to boil
    ⚠️
    Beaker will be hot!
    1. As soon as water begins to boil, turn off the heat and cover the top of beaker with aluminum foil
    image
    image
    1. Leave the egg sitting in the hot water for 11 minutes
    2. As you wait, prepare an ice bath
      1. Use another 1000 mL beaker, fill 1/2 full with water, fill rest full with ice (found in 4th floor ice chests)
    3. After the egg sits for 11 mins, drain out hot water, transfer to ice bath, leave the egg submerged for 15 mins
    image
    1. After ice bath, empty water and place the egg on paper towel
    2. To peel:
      1. Tap egg on counter to break shell into small pieces, carefully peel fractured pieces, keeping egg whites intact
      2. Cut egg in half and scoop out the yellow yolk inside - place to the side in a dish, discard the egg white
    image
    1. Using cooked egg yolk, smash in a dish with a spatula and a small amount of water
      1. Mix until it forms a workable paste
    2. Scoop paste into the red mold and place into freezer for ~2 hrs or until frozen
    image
    image
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