Report 3 - Microbial Techniques Amrita Tharappel Partners:...

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Microbial Techniques Amrita Tharappel Partners: Kito Barrow and Maryanne Mani Section 102 February 6, 2006
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Methods: We used a variety of methods when performing this lab. As soon as we walk into lab we sterilized the bench and our hands to prevent contamination. Then we made the YED media, this was done by adding 3g yeast extract, 6g anhydrous dextrose, 6g agar and 300ml of tap water to a 500 ml flask. After all the ingredients were added, we covered the flask with a small beaker to prevent any particles that are in the air from going inside the flask. We then stirred the flask and put it in the microwave for 2 minutes to sterilize the solution. When opening the microwave again to take the flask out, since it is quite hot, we used paper towels to protect our hands from burning. Giving each group a turn at the microwave since there are two in the class, we stirred the solution around as to dissolve as much as possible, and then put it back in the microwave for 30 seconds. We repeated this process a total of 3 times so that our total time of heating was 3 minutes and 30 seconds. While the YED media was cooling we did the dilution of the yeast culture. First we labeled the for culture tubes B, C, D and E with a pen and then pipette 4.5 ml of sterile water into these four culture tubes. We were provided with culture tube A which had a diluted Yeast culture with OD 600 . In order to perform a successful dilution of 10X, we first removed 0.5ml from tube A, and added it to tube B. Then we capped tube B, and swirled the mixture around so that it would be well mixed. Then we took 0.5 from tube B and added it to tube C, then C to D, and finally D to E, swirling each time after we added the new solution to the culture tube. In order to make this as sterile as possible we used a new pipette for each extraction. The final volumes in each of the culture tubes are as follows A: 0.5ml B: 4.5ml C: 4.5ml D: 4.5ml E: 5.0ml. After performing the 10X serial dilution the YED media was still a little too hot to pour into the plates, so we labeled each of the plates with the following information, our assigned number, the strain name, the dilution tube letter, the date, and we initialed each plate. Each of us then poured the YED media into 4 plates for a total of 12 plates, about 25ml of the media was poured into each plate. We wanted to pour the media as evenly as possible and as slowly in order to prevent air bubbles from forming. If an air bubble did form we were taught how to pop the bubble by quickly running the flame over the YED media. To help keep everything as sterile as possible, we only poured one plate at a time and cover the plate as soon as it was poured and the flask
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This note was uploaded on 09/30/2008 for the course BIOL 2281 taught by Professor Lizhang during the Spring '08 term at University of Texas at Dallas, Richardson.

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Report 3 - Microbial Techniques Amrita Tharappel Partners:...

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