Katherine K. Jacobs: Notebook

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Lab Introduction January 30, 2008

3:30 PM Pipette Use

• Sterilization of the pipette with ethanol
• Pipette #1: 2 microliters blue solution and 5 microliters water
• Set pipette to 7 microliters to test accuracy of previous measurements
• Pipette #2: 20 microliters blue solution and 50 microliters water

Making Solutions:

• Solution #1: KCl Potassium Chloride 0.3 M
• Solution #2: MgCl2 Magnesium Chlroide 0.5 M
• Solution #3: NaCl Sodium Chloride 1 M
• Pour .5 of water amount into container, add salt then add the rest of the water to assist in mixture of two substances
• Preparation of Magnesium Chloride Solution:

(203.31 grams/ mole) x (0.5 moles/Liter) x (0.1 Liter) = 10.1655 grams/ 100 mL

• Preparation of Potassium Chloride Solution:

(74.56 grams/mole) x (0.3 moles/Liter) x (0.1 Liter) = 2.2368 grams/ 100 mL

4: 30 PM Autoclave:

• Mix 25 grams of LB mixture and 15 grams of Bactoagar to a 2 L flask and autoclave for 20 minutes
• Once autoclave was complete, the tape on the flask did not turn black indicating that the mixture was not successfully sterilized.
• Suggestion for solving problem – Autoclave for 40 minutes and set to Sterilize

5:30 PM Making Cell Cultures:

• LB with Ampicillin contains nutrients for the cells to grow
• Keep the culture tubes (with a loose lid) sterile by closing the bag after removing tubes
• The culture should get cloudy while the control should remain clear
• Sterilize the pipette with ethanol
• 2 mL cultures are optimal
• Fill both tubes with the LB first
• Sterilize the loop by placing in ethanol and then flaming
• When removing a colony from a plate, mark on the underside of the plate with a marker which colony has been taken
• Incute the culture overnight at 37 degrees C in a shaking water bath