Thursday, June 23, 2011

What's up graduate student office?

I grew up liquid bacteria cultures last night so I could harvest their plasmids this morning, but alas, my liquid cultures didn't grow overnight. They looked this morning pretty much like they did last night:

I grew four different cultures last night from the same bacteria colony. This colony I has semi-recently used for a plasmid harvest (plasmid prep), so I was shocked to see they hadn't grown. On the left (the darker cultures) is 2xYT buffer (2 times the amount of Yeast Extract Tryptone), an especially nutrient-rich media, and on the right (the lighter cultures) is the standard LB broth (which I just learned from Wikipedia is commonly incorrectly called Luria broth). I set up one of each culture at 1 volume and 2 volumes of the antibiotic ampicillin. I usually grow my cultures at 2 volumes (100µg of amp per milliliter). The amount of antibiotic is very important--let me explain: we're growing these bacteria for their plasmids (sort of like growing people for organs, like in the movie The Island--sorry if I spoiled that one for you hahahaha), and the plasmids have a gene for antibiotic resistance in addition to the other parts of DNA we're cloning the plasmids for (click here for a 101 post on plasmids). Therefore, any bacteria that have a plasmid should be able to survive in the presence of ampicillin. This means we need to add ampicillin to our cultures, to weed out the bacteria that don't have the plasmid. However, if we add too little ampicillin, some bacteria that don't have the resistance gene (from the plasmid) may still be able to survive. If we add too much ampicillin, none of the cells may survive. It's a catch-22: too many bacteria cells (especially those without the plasmid) due to no or too little ampicillin, will give us poor plasmid harvest yields. (This is particularly problematic because bacteria without the plasmid may end up growing faster than counterparts with plasmids, because they don't have to invest the energy into making the plasmids.) However on the other end, too much ampicillin will prevent the growth of cells and I won't get any cells to harvest their plasmids.

In my case, my bacteria colonies were probably too old to start new liquid cultures from. Bacteria colonies are best used if they are actively growing, because they are their healthiest at that point. We'll have to see if I can get these current colonies to grow anymore.

In other news, I'm using my new graduate student desk (which I'm sharing with my labmate Jessica):

It's pretty cool to have a space in the graduate student office, because now I finally feel like a graduate student at Clark. Just settling in as you can see. Jess and I will be using this space primarily to do our reading and writing, but it could also be used to plan out experiments, do research, etc.

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