Cell Culture Preparation of IMD Null Eggs

20220103 Making Yeast Plates

  • Used yeast paste that Kent had made and autoclaved twice. I think this yeast was made the week of 20211220
  • Then that afternoon, made 2 apple juice yeast plates with 2 steaks of yeast on each plate

20220104 Setting up IMD null Flies to Lay

  • Set up 5 vials of IMD null flies to lay on the yeast plate at about 2pm

20220105 Day 1 Cell Culture

  • Collected the apple juice plate at ~9:45am and replaced the cage with a new plate for more laying
  • Followed the Prepping Cell Culture Protocol
  • Used freshly made that day 20% FBS Schneider’s medium, and 0% FBS Schneider’s medium with NO gentamicin:
    • 2 50mL conicals No FBS:
    • 49.5mL Schneider’s Drosophila medium
    • 500ul 100X antibiotics
    • 1 50mL conical 20% FBS:
    • 39.5mL Schneider’s Dosophila medium
    • 10mL FBS
    • 500ul 100X antibiotics
  • Flies laid a good number of eggs, however in one of the last steps the cell pellet seems to have gotten sucked up in a wash or something because the usual cell clump was no longer present when dispensing into the flasks. And there were few cells in the flasks after settling
  • Flasks were left to settle and imaged the next week

20220106 Day 2 Cell Culture

  • Collected the apple juice plate at ~9:45am and disposed of the flies
  • Followed the Prepping Cell Culture Protocol
  • Used the 0% FBS and 20% FBS Schneider’s medium made the day before
  • There were many more eggs on the plate this day
  • Flasks were looked at but not imaged until the next week

20220111 Imaging Flasks

  • Flasks from 20220105 did not have many cells at all:
  • Flasks from 20220106 had a good number of single cells and clumps still looking pretty alive:

20220112 Adding Medium

  • 1mL of 20% FBS medium no gentamicin was added to 1 of the flasks. It was marked as FA