Field of Science

Variant and recombinant competence phenotypes

One of the projects the post-doc has developed is to identify the genetic differences responsible for the very low competence of a clinical strain of H. influenzae.  This strain (86-028NP, 'NP' for short) transforms 100-1000-fold less efficiently than the standard lab strain Rd after the standard competence-inducing treatments.  To identify the genes responsible for this difference he transformed competent Rd cells with DNA from NP, and screened the recombinant cells for ones whose induced competence level had decreased (he had lots of help from a very diligent and competent undergraduate).  (The strains they screened were the same strains whose genomes he sequenced to map their recombination tracts.)  The undergraduate identified one strain whose competence was about tenfold lower than Rd's.

Now I just want to confirm that the recombinant strain does have an intermediate phenotype, and, more generally, to carefully check the phenotypes of both parents and the recombinant, under all of the conditions we know that affect competence.  So I want to check competence at all stages of culture growth in rich medium (a detailed time course) and after transfer to the starvation medium MIV.

Below are the results of a simple time-course experiment using cells growing in rich medium.  It's not a detailed time course since I only did 4 time points, so I plotted transformation frequency as a function of cell density rather than of time, but it clearly shows that the recombinant has an intermediate phenotype.

Next I'll do a better time course and MIV-induced competence.  I should also test induction of competence by transient shift to anaerobic culture, and by addition of cAMP to log-phase cells.

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