Happy Independence Day! (A slightly waylaid post).

Last time I posted, I talked about the US at the World Cup. Unfortunately, Ghana got the best of the team in an intense game that went into extra time. Hats off to a team that played an incredible game.

Luckily for me my project gets to progress to the next stage. Finally the control plasmid that contains all the mutations and has the temperature sensitive (Ts) phenotype was found in our stock and sequenced verifying the presence of the five amino acid changes. This is important that we have a control for the next phase of testing.

A new primer was also ordered for the one mutation that doesn’t seem to integrate when not in combination with other primer sets. This primer has come in and will be tested in the next few days.

The next phase of testing involves electroporating mutant plasmid DNA into Burkholderia thailandensis. Electrocompetent cells are grown up and plasmid DNA is electroporated into the cells. The first trial consisted of one control (the fully mutated temperature sensitive plasmid) and the wild type plasmid with one amino acid mutation (R19C). Luckily, or unluckily, for me this worked too well and the dilutions were too high. I had too many colonies to count. We now know that the electroporation works now it is just a matter of dilutions to obtain isolated colonies to patch and test for Ts.

I apologize for the sporadic posting but it mirrors the way of research, every once in a while things go right. As my uncle loves to say, “even a blind squirrel finds a nut every once in a while.”

As always, thank you for reading and go in peace.

Erin

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