Today we are making e.coli for our DDW experiments. The first step is to make the e.coli medium. We made up a small batch of LB Broth, using the instructions from the Sigma-Aldrich website. We only made 100mL of LB Broth, instead of 1L, as follows:
- Mixed 2 g of LB Broth powder with 100mL of distilled water.
- Autoclave for 15 minutes at 121°C.
After the mixture cooled, we measured 10mL into two test tubes and mixed in a minute amount of e.coli.
Next we placed the test tubes in the New Brunswick Scientific Innova 4300 Incubator Shaker at 37° C. Our shaker’s thermostat is about .3°C – .4°C high , so we stuck a thermometer probe in the shaker to get an accurate temperature measurement.
We set the incubator shaker at 70 RPM for a few hours. I have to leave for class, and Anthony wasn’t sure if we needed to leave it longer or maybe even overnight. Please check his blog for the final details.
Feb 07, 2012 @ 17:40:39
It is important to note that the reason we used two test tubes is because we’re growing two different kinds of e. coli. One is from Stratagene and contains the pBluescript vector, and the other is from DNA 2.0 and contains the pALS (made by me) vector. The pALS e. coli is less likely to work because it has been sitting in a stab at 4C for about 12 months.
Feb 07, 2012 @ 21:08:46
I checked the e coli before my GPSA meeting and nothing had developed yet so I decided to leave it for overnight growth. I’ll check it out in the morning.
Feb 08, 2012 @ 10:52:43
Yay, we have e. coli growth in one of the tubes. I’ll work on the nanodrop until you come in and then we’ll figure out what we have.
Beginning E. coli growth | IheartAnthony's Research
Feb 08, 2012 @ 17:00:12