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From 2010.igem.org

9/2/10

Results from streaking – Transformed DNA grew on all four plates and progressively grew less when the Amp Con. was raised. So selection was possibly made. We will run a Electro gel to look for possibly the bands need. Electro comp cells won’t grow in the presence of Amp without transformed DNA. Looking at Results when the Con. of Amp is too high you get limited GFP expression and I believe that electro are contaminated by me after/when I streaked my plates. Today I will suspend the Transformed DNA into 10 mL Broth with 10 mL Amp and shake.

1) Set up a Clean environment by cleaning table, tools, etc. with disinfectants. then prof. T. told me to do it in Bio hood So I did :)

2)Moved all into bio-fume hood

3) Placed 10 mL LB Broth into Sterile 15 mL Centrifuge tube then from streaked plate #3 I pulled a Colony from this. A Colony is placed for resuspendion in this. Then I added using a steril 5 mL pipette I added 1 mL LB/Broth into 6 sterile 1 mL Centrifuge tubes. then added a Colony to each. then using my .5 – 10 mL micropippettor I put 1.0 mL of Amp in each 1 mL Centrifuge tube.

4) I added a large Colony into 10 mL Centrifuge tube w/LB Broth add 10 mL Amp using my 2-20 mL micropipettor.

5) I have placed 10 mL tube into incubator until tomorrow. Extraction of DNA of KBBa _K131010 from Electrotransformation Irene has performed all operations of this protocol and will finish by placing spl. in the box freezer in back room -20 C

9/2/10

Protocol For Making LB-Broth/Agar Today I made 2 L LB/Agar I placed

1) 10 g LB-Lennox into 4 – 1000 mL capped Elenmyier Flasks

2) then placed/weighed 7.1 g Molecular Biology Agar into the same(4) Elenmyier Flasks

3) added 50 mL 18 ohm water into each of the 4 Elenmyier Flasks

4) then microwave on Beverage 8 oz. setting 6 times to melt /dissolve Agar into solution.

After dissolved I placed into Autoclave on Liquids Setting and let run overnight. note Irene made 250 mL and placed into Autoclave. I removed after done labeled 9/2/10 J.H. LB/Agar. and put in fridge in back.

9/2/10

I wasn’t able to fit all 4 – 1000 mL capped Elenmyier Flasks into Autoclave at one time so I placed the 2 in autoclave. and 2 in a 40 C Hot water bath. The 2 in Autoclave we placed inside pan and set to Liquid setting and started which is in 121 C at 15 min. will autoclave the other 2 Friday 9/3/10 9/2/10 J.H.