Using the realtime PCR machine to measure GFP expression

As it turns out, the settings for reading the SYBR green dye also works for fluorescing GFP meaning that the realtime PCR machine in McM221 can be used in this way. Tried a quick test to see if I could get it to work. 4 samples monitored over the course of an hour at 37 deg:

• 30 ul LB broth uninoculated
• 30 ul LB + E.coli HB101 
• 30 ul LB + E.coli HB101 containing the pGLO plasmid, harvested off of an LB+Amp+Ara plate
• 30 ul LB + E.coli HB101 containing the pGLO plasmid, harvested off of an LB+Amp+Ara plate, but then also amended with 5 ul of 0.4 mg/ml arabinose. 
• The A580 reading for the E.coli without the plasmid was 0.29 which translates roughly into 1E8 cells/ml
• The A580 reading for the E.coli with the plasmid was 0.33 which translates roughly into 1.1E8 cells/ml
• Made a program to hold the temperature at 37 deg C and then to take fluorescence readings every minute for an hour.

Things to note from the graph

• bacteria without pGLO showed same level of fluorescence as just the media
• bacteria with pGLO were consistently higher
• bacteria without pGLO both showed  steady levels of signal across an hour while bacteria with pGLO increased. My assumption is that since the bacteria with the plasmid came from a plate that already had arabinose on it, the bacteria already were expressing GFP. Increases across time indicate the increase in GFP - could be due to cell doubling or due to increased GFP expression. Either way the pGLO cells without arabinose shouldn't have shown an increase. 

At the end of the day this shows promise as a tool for us to develop labs around, and of course research too. 

tags

Technical note - just figured out how to appropriately tag my blogs and display the tag cloud (see to the right) so now you can click on the tag of interest to pull up any posts related to that. Cheers

Straightforward video conferencing with iChat/Messages on Mac computers

Straightforward video conferencing with iChat/Messages on Mac computers

A few years ago I was playing around with the iChat Theater mode as a tool for easy off-site presenting. The cool thing about Theater is that it not only allows you to show someone a series of slides or images that you have, it also keeps you in the picture to narrate it. See the pictures below when I recently tested it. The first picture is what the person on the other end saw - me in the big picture and him (which happens to be me too) in the small picture. Next I dragged my pdf file into the main window on my computer - this became the main picture and my face shrunk down to the lower corner (you can see this in the lower picture) So now I can show the other person (or people) my presentation/pictures/videos/etc while they can still see me narrating.

Did this with a winter term class a few years ago - had a colleague video in and show us some of his research slides that related to a paper we had read. He was projected on the screen and so he filled up the whole screen. When he switched to Theater mode the transition I described earlier happened and that was a "wow" moment for the students that they continued to talk about for quite some time. I wish the wow had been for the material, but I'll take it anyway....

I recently upgraded to Mac OsX 10.8 which moves iChat to Messages. It's not as straightforward to do this anymore but it's there.

preparing prelab/lecture material video

There is a lot of talk about classroom flipping these days and I can see where this will be great - get the content deliver "out of the way" before the class meets, then spend the class time applying the content and trying to get it to a level of working understanding. In many ways we've been doing this in our Biology labs already via prelab content on video that is then applied with hands-on activities during the lab. One of the key things you need to be able to do easily is to capture your lecture/prelab/etc.  Dan Reis at TLT showed me a great tool the other day (free too) called Screencast-o-Matic. SOM allows you to identify a portion of your desktop that you want to be filmed. It also can simultaneously capture video of you so that the students can see you giving the talk.

Here's a screenshot of it in action. The dotted line is what will be captured and it will embed the video of you in there if you want it to. 

SOM has export functions for exporting into mp4, flv, and avi formats. It can also export directly to YouTube which would have some advantages. 

Note that while there are ways to do this kind of thing without the video, inclusion of the video I think is a nice touch that students will appreciate. 

http://www.screencast-o-matic.com