Category Archives: Ecoli1

E. coli growth experimental setup and data (on FigShare)

E. Coli Growth over 4 hours. Anthony Salvagno, Alexandria Haddad. Figshare.
Retrieved 20:17, April 20, 2012
hdl.handle.net/10779/b627469dabcd4034053cc53040d4dcbd

I went through the data that I posted on Tuesday and realized it was even less useful to people than I expected. I almost didn’t even know what I was looking at! Anyways, I did a couple of plots in excel with the data (which can be found on FigShare along with both the original data and the revised and cleaned data) and tried to extrapolate some other information. But first let’s discuss the experimental setup.

So on Monday, Alex created a starter culture from the E. coli we grew on plates last week. Then on Tuesday (I realize how not very real-time this post is for me, but the data came out in real-time which is also important) we made 3 dilutions of the starter culture to track the growth of the E. coli over time. We did:

  • 1ml in 9ml of LB broth (1:10)
  • 2m in 8ml of LB broth (1:5)
  • 5ml in 5ml of LB broth (1:2)

Every hour we took an absorbance reading from the nanodrop and read the 600nm value (A600 according to the machine). We also reblanked every hour according to the instructions from the nanodrop. The initial readings were:

  • starter culture – 1.076
  • 1:10 – 0.097
  • 1:5 – 0.23
  • 1:2 – 0.665

It is interesting to note (and I literally just noticed this), that the initial readings are almost exactly what the dilutions are, ie 0.097 is ~1/10 of 1.076. Good for us!

In the FigShare data, you will find the original data (which I linked to in my post on Tues, but in Google Docs instead of Excel) and a revised data set. The data is messy but the graphs are interesting.

Also I tried to link the 3 data sets together into one coherent graph, but the time series doesn’t seem to match up right, or maybe it does and I just think it doesn’t. The 1:5 dilution seems to provide a bridge between the data in the 1:10 dilution and the 1:2 dilution. After about 2 hours the 1:10 sample overlaps with the 1:5 and likewise the 1:5 begins to overlap with the 1:2. Also at hour 3, the 1:10 sample overlaps with the 1:2 sample.

Because of this I tried to graph the data as one continuous set. It seems Like it may be alright, but I feel that the in the 1:2 sample there isn’t much growth in the 4 hours, which is reflected in the 3 data set plot, but it doesn’t look like it peaks in the continuous graph. Hmmm… Anyways check out the FigShare data.

PS I’m including the two plots I’m referring to below.

 

Yeast and E. coli Day 2

First off, how come yeast has a commonĀ  name but E. coli doesn’t? Can we call it shitus?

Anyways yesterday Alex and I did some follow up work after starting some cultures. She did a good job notebooking the experience so I won’t double up on her thoughts. Check that out here.

We took some pictures of the e.coli and the yeast which turned out pretty bad. I’ll have to try and take some better images today or something. In general though it looks like the yeast we have isn’t what we thought it was. It may not be a mutant of S. cerevisiae but may be some version of Schizosaccaramyces pombe (I hope I spelled that right, I like the first name a lot) because it looked nothing like budding yeast.

The result of this is that I will be ordering some new yeast straight up from atcc.org that is of some variety that I commented on in Alex’s notebook (hopefully).

E. Coli and Yeast Incubation

I’m showing Alex some sterile technique and how to inoculate yeast and e. coli via liquid media and solid (agar) media.

Today we plucked a few colonies of yeast from our starter batch (which may need to be replenished soon) and streaked the cells onto some YPD plates. One plate is in an incubator/shaker at 24C and the other is sitting at RT. We also inoculated a colony in 10mL of YPD.

For the E. coli, we did essentially the same thing, but we are doing 1 plate of LB media (agar) and 1 10mL test tube of LB broth.

Tomorrow we will try for some OD readings, and then we will try to setup time point growth experiments. It should be noted this is all in regular H2O. Right now I want Alex to get familiar with the process and then we will start doing some DDW and D2O studies probably in a couple of weeks.

#SciFund Challenge Accepted!

If you watch “How I Met Your Mother” you’ll get the title of this post. But if you don’t then I’ll let it be known that I have just completed my submission for the second #SciFund challenge which is hosted by RocketHub.com. Here is my submission:


Effects of DDW on Microorganisms

Yeast: Day 2

So my overnight culture grew, that’s cool. The E coli did not though. That’s not. To make matters worse, the -80C freezer had some weird malfunction and it was nearly RT in there. Hopefully nothing is ruined. I’ll restart experiments on Monday and make new batches of yeast and e coli spreads and liquid media as well. Then I’ll make some glycerol stocks so that I can be ready for all eternity.

On top of all that I’ll reacquaint myself with doing transformations and purifying plasmids because there are some vectors that I need to replenish. Not for the purposes of these experiments, but just because.

I’ll try and do some preplanning over the weekend and through Monday.