Monday, August 28, 2006

Been a while...

And, sadly, after an exhausting day I'm not going to post much to make up for it. Tomorrow is dedicated to composing pieces of our sxy manuscript, including: crunching today's data, getting figures together, writing my and Laura's Material and Methods sections, finishing the strain list, and (time premitting, which I doubt) thinking about elements of the discussion.

Now, a little bit of thinking on MurE hypercompetence mutants: Inspired yesterday by handing Rosie a figure showing that addition of AMP or GMP to MIV cultures results in reduced Sxy production. A neat feature of the data is that AMP has a more dramatic effect than does GMP on Sxy levels, and this is exactly what we see in terms of repression of transformation frequency. It got me to thinking that I saw the same thing when treating murE749 cells with nucleotides, which got me wondering if murE749 induces competence through the standard Sxy + CRP mechanism, does it do so by specifically inducing sxy? Caixia describes testing one of Laura's sxy::lacZ fusions in the murE749 background, but the strain described in that paper is misslabeled in the paper's strain list (ie. our records show the strain number from the paper to correspond to a xyl mutant of some sort). I sifted through Caixia's notebook and could find most of the other murE experiments, but not this one... So I don't know what type of fusion Caixia tested: protein or operon? It's possible she tested an operon fusion, which we now know doesn't acurately reflect Sxy abundance in the cell. I could directly measure Sxy levels in the murE749 strain, and if we found that they were increased, we could be fairly confident that murE749 induces competence through Sxy. This would be interesting, but also just another piece of the complex and unresolved story of how sxy is regulated.


At 7:08 AM, Blogger Rosie Redfield said...

Jessica did one or two microarrays comparing the murE749 mutant to wildtype, I think in early log. And I think she found that mRNA levels of sxy and of all the CRP-S genes were up.

We really need to get access to GeneSpring so we can recheck this and other array results. New murE results would be great.

At 7:14 PM, Blogger Heather Maughan said...

I recommend a statistics package I used for analyzing my microarray data. It is free and does better statistics than GeneSpring (or so I hear). It is implemented in R, which has a steep learning curve, but is also the best for stats.


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