Counting cells and taking names

I spent a good portion of Friday morning and early afternoon in lab doing some work and figuring out where to go next in my project. I talked about different kinds of reporter plasmids we may use in this project with my professor (we may incorporate other genes that have plasmids already made) in addition to what kind of antibiotics we're going to use.

Two types of antibiotics have been used in recent projects transforming diatoms: nourseothricin and zeocin. The former is 100 times less toxic than the former, so we're going to start with using that.  Right now we're growing the plasmids without any selection pressures (no antibiotics). After we get a feel for how well the cells grow on the agar plates, we'll test them with nourseothricin. First of all, we need to make sure nourseothricin kills our diatom cells. This way once we transform them with our plasmids (which will include the anti-nourseothricin plasmid, nat), we'll know that only diatoms that have been transformed with nat will survive when exposed to nourseothricin.

In order to grow the diatom culture on an agar plate, I had to count them first. This entails pipetting a small amount of diatom liquid culture onto a hemocytometer, which is essentially a microscope slide with gridlines on it. The hemocytometer allows us to estimate the number of cells per milliliter. That way we can know about how many diatom cells we're putting into a fresh culture or agar plate. I'll have to get a hold of a special camera or something so I can take pictures of what I see on the microscope!

Once I counted my cells, I plated four different amounts on four different plates. This way, we can see how different starting amounts of cells affects how they grow. These cultures will take up to a week or so to grow, so I'll have to find something else to keep myself occupied in the lab in the meantime! :-p

After they've grown up a bit, I'll start testing the nourseothricin on the cells like I mentioned above. We could do this by soaking paper discs (about the size of hole punches) with antibiotics, or spreading antibiotics on a plate and then growing them.


I talk about all of this stuff in my video blog post below:



Outside of the lab, in the so called real world, we've been having a slight thaw. This has been warmly welcomed on campus. It rained most of the day yesterday and today it is in the mid 30's. I was wondering to myself the other day what would happen if temperatures jumped up into the 60's for two or three days. All of this snow... would lead to massive flooding. How awesome would that be!?