Day 1

7 a.m. Get ready for work and catch the 7:35 a.m. shuttle from Parnassus to Mission Bay...arrive at the shuttle stop a bit early. Splurge on a Frappaccino at Starbucks. I try not to waste money on coffee...after all, as a grad student, $$ can be tight.

8-11 a.m. Set up destains of gels (protein gels...this is a method by which we can separate proteins based on size. It is an analytical method where the proteins are separated by mass on a small clear gel. Each protein resolves into an individual band. When we are done running the gels we can stain them with a blue dye that stains the proteins and the rest of the gels. In order to remove the stain from the rest of the gel (but not the bands) we place the gels in destain. It smells like methanol and vinegar. Gross...).

Place western blot in blocking solution. After running a gel you can transfer the proteins onto a piece of white paperlike membrane. This membrane can then be probed with antibodies specific to the protein of interest, helping me to determine if my protein of interest is in the sample I ran on the protein gel. The western blot process is long, involving frequent buffer changes.

Check email. Take care of some administrative stuff regarding who will pay my stipend this year.

Start to work on some slides for a presentation my PI (principal investigator) has to give on Thursday. He asked for the slides last Wednesday but I was out of town Thursday-Saturday for our departmental retreat. I'll be putting together some of my data and some background on PowerPoint slides.

Mornings are my most productive time...not many other people are in the lab yet so I can get things done without much distraction.

Head into the Tissue Culture room to take care of some mammalian cell lines. Unlike bacteria and yeast, mammalian cells are usually taken care of with extreme care and caution to maintain sterility. We have a separate room for tissue culture. Working in the Tissue Culture room is often monotonous (lots of pipetting things up and down, over and over again) and lonely. Since we only have one hood (part of maintaining sterility) in our TC room, only one person can work there at a time. I spend about one hour or so maintaining my cells and splitting them (so that they don't get too crowded in their plastic Petri dishes).

11:30 a.m.-1:15 p.m. Group meeting. Our lab meets every week on Tuesdays to discuss our work. Each week, one person presents. I present my work about once every four months. It is a forum for discussion and practicing our presentation skills.

1:15-1:35 p.m. Scarf down lunch.

1:35-6 p.m. More busywork around the lab. Repeating experiments I've done in the past trying to get them to work. There is a LOT of repetition in graduate school. In between I have brief moments during downtime in my experiments to chat with other folks in the lab. We discuss our work, recent papers and ideas for future work. I particularly enjoy talking with my baymate, another graduate student in the lab. He's a few years ahead of me and has incredible insight into his own work and the work of others. We try to troubleshoot some of the problems in my experiments and his.

I also find some time to update a slide for my PI's talks. An interesting paper came out recently and, accordingly, I need to update a domain diagram to show the discovery of a new domain in one of the proteins we study. This domain was not predicted by sequence homology (the sequence homology is too weak); instead, a crystal structure showed that though the amino acid sequence is fairly divergent, the protein fold is very similar to that of a known domain found in proteins involved in cholesterol metabolism.

6-6:40 p.m. Set up some experiments and plan a schedule for tomorrow. Of course, you can make a plan detailed down to the hour, but invariably, one step will fail or have problems, thereby throwing the entire schedule off. That said, I still like to have a list of things that I would like to accomplish.

6:40 p.m. Catch the shuttle home.

7:10ish p.m. Arrive at Parnassus and walk home. Meet with some friends and go to the gym. Grab a burrito after working out at the climbing gym and head home. Shower and get ready for tomorrow.

Day 2

6:30 a.m. Arrive at the lab. It's a VERY early morning. I have tons of things to do. As I'm sitting at my bench doing a gel extraction. (I cut two different pieces of DNA into pieces and am going to ligate them together to make a new DNA construct. Before I can do the ligation I need to get rid of the digesting enzymes and uncut DNA. To do this I run the DNA on a horizontal Jell-o-like agarose gel. I then stain the gel and cut out the bands of interest and put them into a little 1.5 ml plastic tube). I notice the bay that I'm in start to glow orange. The sun is coming up. After doing the gel extraction I set up a ligation reaction (and all the appropriate controls. It is interesting how sometimes you can have four-five times as many controls as you do experimental reactions!). I then transform the ligated DNA into competent bacteria (basically these are bacteria that have been treated with chemicals to poke little holes in the membranes so that DNA can enter) and place the bacteria to shake at 37 degrees (Celsius, we never use Fahrenheit in science) to "outgrow" for an hour. I put petri dishes with growth medium and a selective antibiotic in the other 37 incubator (not shaking) to dry. In between all of these things I go and check on my mammalian cells and set them up for an experiment tomorrow. They are growing well and aren't too crowded yet. After I plate the cells (place the cells in a liquid suspension onto the dish and spread them out with the sterile glass beads) I pop down to the White Star Café and get a bite to eat. Another graduate student comes with me and we discuss protease inhibitor design.

10 a.m. The lab is getting much more full. I spend some time reviewing the logic of my experimentation and the biochemistry behind it. I also take this opportunity to catch up on reading papers. I scan over a paper that I'm supposed to present at a journal club. I've already read it a couple of times, but now am trying to figure out how to present the information in a concise and interesting way.

11:30 a.m. There is another set of slides I have to do for my PI. While working at the computer, I glance out from time to time at the construction workers and note how different their jobs are from mine. After all, they can SEE their progress at the end of a day. Science moves at a pace that is glacial compared to that of construction work. Keeping motivation high and working through difficult times/problems with no end in sight is one of the KEY aspects of graduate education. Also, because we all tend to work on our own projects, it can get lonely from time to time.

1 p.m. Lunch -- brought it from home. Not a lot of places in the vicinity to eat. And it wastes too much time.

2-6 p.m. Set up a cell lysis assay for tomorrow. In this assay I treat cells with the protein I study and look to see a cell death phenotype. I came upon this effect by accident but feel that it may help me figure out the role of my protein in biology. The only tough thing about this experiment is that I have to be around to count my cells every hour for several hours. The cells are grown ahead of time, counted and then plated.

6:30 p.m. Go out for dinner with a friend.

Day 3

10 a.m. Arrive at the lab. Set up some molecular biology experiments and talk with a post-doc in the lab who has a similar project to mine about the general direction of our research.

Lunch (go out to 18th and Connecticut to a Thai restaurant with some lab friends). We discuss the new post-doc candidate and possible projects for her.

After lunch, I sit down and plan a purification scheme for one of the newer proteins I'm studying (I'll do the purification tomorrow or the day after...once I have enough hope and motivation to face failure again).

2:30 p.m.-2 a.m. Time to do the lysis assay. I wash the cells (mammalian cells that I set up yesterday) in cold PBS (buffer) and get ready to add protease. I carefully recheck my calculations and add protease to some of the cells and not to other samples. This way I have a negative control (it is really important to control your experiments well). I then take some microscope pictures and count cells at timepoint zero. Now all that is left to do is to count cells every hour. It takes about 30 minutes to prepare and count the cells for each timepoint so I have 30 minutes of downtime every hour. In this time I read papers in between timepoints. By the LATE night I get tired of reading papers and just listen to music and read a novel. Drink some more coffee. Of course, my data from this experiment didn't work out as planned (because all the cells detached so I couldn't keep track of the assay).