To Do
- Camera stuff
- Kinesin activity assay?
Camera
- Koch wants to use the camera as a gel camera.
- He wonders how well it works wired
- The answer to that is very well
- The camera can dump all the photos it has taken with the push of one button
- The destination folder on Larry's computer is in the C: drive
- It also sub categorizes the photos by date taken
- As for a long usb cable, Im pretty sure they make em almost as long as you would ever want
- The camera uses usb mini-b, so we may want to get an adapter (the thought being get a really long usb A cord and get an A to mini b adaptor, that way the long cord can be used for other things as well)
- Amazon has usb A extension cables (15-20ft) for under $10
- So Koch ordered two Canon Cameras and wants me to try to get the second one setup
Kinesin Activity Assay
- Brigette wanted to try to do the phosphorous assay kit that we got from cytoskeleton
- From what I know about this, we mix up tubes and ATP and let it sit at different times and measure the phosphorus levels
- We then mix our kinesin and the tubes and ATP, let sit at diff times, and re measure the phosphorus
- Right now, we are going to polymerize some non-labeled tubes
- Brigette is writing up some procedures which I hope to copy from her and paste in my notebook
Procedure
- Taking note of what we did as we do it:
- 1 aliquot of the unlabeled tubes
- They are in the -80c in the large falcon tubes
- Polymerize them for 20min @ 37C
- Add 99ul of BRB80T for a total of 100 ul of tubulin
- note: the BRB80T is cold in the 4c fridge, so i warmed the tube by hand for ~5mins
- Tubes + brb80t created at 12:10pm
- Brigette is making a little bit of everything to check and see if any of it ends up killing the kinesin
- We are doing just tubes, just kinesin, tubs + kinesin, the motillity assay with casein, etc
- The idea here is to make different kinds of samples and let them all sit for about the same amount of time (~30mins)
- Samples to be made:
- 30ul MT + 30ul Kin
- 30ul MT + 30ul Kin + .6ul ATP (1uM ATP)
- 30ul MT + 30ul BRB80T + .6ul ATP (1uM ATP)
- 30ul MT + 30ul Kin + 1.2ul ATP (2uM ATP)
- 30ul MT + 30ul BRB80T + 1.2ul ATP (2uM ATP)
- Note: When i say MT I mean MT+AF (motillity soln w/o casein)
- Second Note: some drops got stuck to the side walls of the tubes, so Brigette washed them down with a couple ul of BRB80
- We let the samples sit for ~10mins to allow the reactions to complete (since adding the dye and quencher stop the reactions all together)
- Turns out it is over diluted and Brigette didn't get any useful readings from it
Trial 2
- We are redoing it, but w/o any naked brb80
- Samples:
- Motility Assay alone
- Mot +1uM ATP
- Mot +2 ATP
- Mot+ 3 ATP
- Mot
- Mot+Kin
- Mot+kin+ATP
- Mot+kin+2ATP
- Mot+Kin+3Atp
- Allowing it to sit and react for ~ 30mins
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