Huntingtin sample preparation before the holidays

Before the university shuts down for the holidays, I have been busy in the lab making huntingtin protein samples. I purified wild-type huntingtin with a polyQ stretch of Q23 as well as disease versions of the protein with Q54 and Q145. Whilst Q23 huntingtin protein can be trivially expressed and purified, the longer polyQ length versions of this protein give much lower yields and there was hardly any Q145 following a gel filtration purification step.

I also made some samples for cryo-EM which are fixed with glutaraldehyde. In order to identify the 2 ends of the protein, I used N and C FLAG-tagged versions of full-length huntingtin with FLAG antibody to generate 2 terminally tagged samples. I have sent these off to my collaborator Susan Lea to make grids and screen the samples over the holidays – fingers crossed we get some nice data!

Gradient fixed (Grafix) huntingtin samples – a clear band of huntingtin is seen in fractions 7 and 8 for each run of this sample.

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