Wednesday, August 22, 2012

Follow the biology (pt. 3), slight end of summer update

I'd like to apologize first about how sparse my postings have been, but, starting this blog just happened to coincide with the summer conference season as well as the start of the semester here at U of A.

I thought I'd jump back into this by giving a very brief update on an open science project that I started to talk about a few weeks ago (parts. 1 here and 2 here). Over the summer I had an undergrad working on the project in order to isolate transposon mutants that don't display the brown phenotype. After screening ~1000 independent colonies, I'm sad to say that every single plate looked exactly like this:

There were always a couple of empty wells where the colonies didn't grow but, eventually, every well turned brown. Astute readers may point out that well A4 doesn't look so brown (hence why I have this picture lying around). However, even this well turned brown after an additional day of growth....and every single colony isolated from that well (~20 or so) turned brown so the late browning wasn't just contamination. Of course, it's possible that whatever pathway leads to the brown phenotype is essential for cellular growth so that mutations would never be identified from a transposon screen. 1000 colonies isn't enough to make a case either way, but it's enough to begin thinking about the next step.

Now that the summer is over, the talented undergrad working on that project has moved on to grad school in another department so I hesitate to devote much more time to the transposon hunt. While it's possible that I could eventually isolate a non-brown colony, there are better uses of my time right now. It was worth a shot but this is a point in my own brain where I cut bait and try a different strategy (especially when the undergrad has moved on to another department). So now I'm going to isolate some genomic DNA from this mutant and sequence the genome in order to identify any potential genotypic differences. We'll see how that goes.


  1. I posted a suggestion - where did it go?

  2. OK, that one worked (nasty tricky captcha). My suggestion: Use pooled DNA from the transposon mutants to transform the wild-type strain to antibiotic resistance and screen for the brown phenotype. Now you have a transposon that's linked to the gene causing brownness. If you don't get any brown transformants it's probably not a single-gene phenotype.

  3. Thanks Rosie! I thought about that in passing, but have only done it with phage (and wasn't quite sure of how closely linked I could get the marker). Will definitely try now!

  4. I tried to post a suggestion earlier, it disappeared. I vaguely remember that the dark brown phenotype in P aeru is caused by pyomelanin synthesis. Dianne Newman's group published a paper on finding genes involved, that might help you in your quest (AEM, I think).

  5. Should have checked! it was JBact
    Good luck!


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