This is a bit of an aside to the posts about my research, but it is what I have been doing the past few days, so I thought I would write about it. I am eventually going to end up doing research with bacterial flagella, so we need a way to get purified flagella from bacteria. For reference, this is a bacteria with three flagella:
So, basically, we want to grow bacteria that have these flagella, separate the two, and just take the flagella. Easy, right?
The first thing we need to do is grow our bacteria. The kind of bacteria pictured above is the same kind we use, E. coli. The way you grow these bad boys is you first take some that you either bought, or grew previously out of the freezer and spread it out on a petri dish that you put bacteria food in. The bacteria food in our case is some concoction called LB Media, but it is essentially just a bacteria Outback Steakhouse.
Once we have grown some bacteria in a petri dish, we take one little colony of that bacteria and put it into a liquid form of the bacteria food. Once it is in the liquid food, those few bacteria we put into the liquid divide and grow in number quite rapidly. Pretty soon, we will have a big beaker full of liquid that is filled with bacteria. The next step is to separate the bacteria from the liquid. To do this, we put the mixture into a piece of equipment called a centrifuge. All it does is it spins around very very fast. Things that are dense will sink to the bottom and lighter things will come to the top. In our case, the bacteria will be forced down to the bottom of the container because they are denser than the liquid food. So, when our sample comes out of the centrifuge, we just get rid of all the fluid because the bacteria will have been forced down to the bottom of the container and formed a pellet that will not pour out.
However, these bacteria, when they come out of the centrifuge, still have their flagella attached. Now, we need to figure out a way to separate these little guys from their flagella. It turns out that all it takes to separate the two is just a vigorous shaking for several minutes. So, we add a different fluid to the container with the bacteria pellet so that it dissolves the bacteria into solution again. After some shaking, we have the two separated from each other, but they are freely floating around in this new container. We need to separate the bacteria bodies from their flagella. To do this, we just put the whole mixture back into the centrifuge. The cells are denser than the flagella, so they form a pellet in the bottom of the container that we can just discard.
So now we have a solution with flagella floating around in it, but it is very dilute. We want to make a much higher concentrated sample of flagella. To do this, we make one more trip to the centrifuge. This time, the flagella are the dense thing in the solution, so they will form a pellet in the bottom of the container. We can dissolve this in a small amount of fluid and, Voila! A concentrated solution of flagella!
Another time I will go over ways in which we use these flagella once we have them. If you have questions about any of the steps, leave a comment!
UPDATE: It turns out we use Salmonella bacteria rather than E. coli because they provide more flagella. It is the exact same procedure for both however. Just for reference, here is a picture of Salmonella. It is clear to see that it provides more flagella than the E. coli.