UW Stout/D2O SP22

Introduction

D2O also known as Heavy Water is a form of water that is deuterium. It is heavier than normal H2O and can have many physical and chemical properties that are different than normal H2O. It is known to slow down neutrons and, in some cases, possibly absorb these neutrons. Although it is similar to water it can only be somewhat safely consumed in very small quantities. The experimentation process uses a Wild-Type yeast cell as a control and 9 Knockout Strains for the experimentation. With this, we will observe and record the growth curves of these cells using Optical Density 600 in a 24-hour observation period which in return will tell us the number of cells in each well plate.

Materials and Amounts

• 1ml D2O
• Sterile Water
• Wild-type yeast in 2x synthetic complete media at an OD600 of 0.1-0.2
• Corning COSTAR 96-well clear flat-bottom assay plate
• 9 Knockout Yeast Strains

Equipment

• Biosafety Cabinet
• Micro-pipetting tips
• Micropipettes
• Molecular Devices Spectra Max Plus 384 Microplate Reader

Safety

• Safety Glasses
• Lab Jacket
• Gloves
• Safety Data Sheet for D2O

Procedure

1. Diluting the D2O with sterile water, by following the dilution rates below

1. 1. 50% dilutions (25ul D2O to 25ul H2O) (10 samples of this dilution)(Used only in Pilot)
   2. 0% dilutions (50ul H2O) (10 samples of this dilution)(Control)
   3. 75% dilutions (37.5ul D2O with 12.5 ul of H2O)(Used only in Pilot)
   4. 35% Dilutions (17.5ul D2O with 32.5ul of H2O)(Final Dilution)
   5. 20% dilutions (10ul of D2O with 40ul of H2O)(Used only in Pilot)