Grad Diary 5/10/10

And I'm back!

Sorry for the long wait. As my last post indicated, I was spending quite some time reading the literature and developing my ideas. A month later, I clocked in about five weeks of reading or so and burned through about 50-60 papers (I wish I was exaggerating). It was intense, both boring and exciting, and totally worth it. These periods of intense canvassing through the literature are really critical at times, especially at the beginnings of projects or when things start to become ambiguous. It is a great opportunity to see what is known in the field, get ideas of how things have been done and possibly adapt approaches for new questions, etc. I knew I was coming to an end of my reading when I recognized many of the references listed in the papers I was reading (I call it coming "full circle").

Of course, the reading never really ends. There's always new findings to stay on top of, controversial claims to think critically about, etc, but now I have a good idea of what is known about HPV integration, what isn't and the questions I am interested in asking.

During that time I also got a brand new MacBook, which has completely revolutionized my life, ha ha. My old laptop has been with me since I started college, and it served me faithfully through the past five years. It was starting to show its age, though, and it was time for an upgrade. It's amazing how having access to a sleek, fast and portable computer can really change your work output. I also came to appreciate how much of my research requires or is enhanced by having a computer. Granted, my experiments are done at the bench, but my literature searches, my presentations, data management, etc are all done on the computer, and it's much more enjoyable now that I have a really efficient one at my disposable. The transition from Dell to Mac has actually been kind of fun, mainly because I enjoy learning way too much. And the trackpad is simply too awesome.

I digress. Another big thing that has happened since my last post is the end of spring classes (thank god!). I ended up doing really well. I was especially happy with my success in Immunology, a topic for which I had very little background but which I found extremely interesting and complicated. The end of classes means I can now focus my entire day on research, and when I go home in the evening, I can devote almost my entire time just relaxing and doing things for me, without having to worry about reading or studying for class. It's pretty nice!

Anotherl big thing was that Luis, the senior grad student in Una's lab, gave his thesis defense mid-April, and (of course) passed. We had a nice little reception afterwords and will have another sending-off party at the end of May when he finally moves out. He is going to be doing a post-doc at Yale. He will be greatly missed.

Finally, one of our post-docs, Sunny, finished her time with us and returned to China last week. It was very sad, but she may be returning to work with us again after two years or so (it's weird considering the fact that I will still be here, ha ha). We had a nice little lab outing to send her off and gave her various Penn souvenirs. She's got two papers in the works that we will probably finish via e-mail over the coming months, so I'm sure we'll stay in contact for a bit.

That's all the major news from me for now. I've developed some good ideas for experiments to do and questions to address regarding HPV integration and am now getting started with them. It feels good to be doing experiments again. After things develop for a bit Jian and I will probably start working on developing a secondary project as well. It's always good (especially in the beginning) to work on two or even three things at once, because you never know which one(s) will take off and which one(s) will tank. Either way, the summer looks to be a very productive and fun time.

Grad Diary 4/5/10

Wow...I feel like my head's about to explode.

My project came to an end a little over a week ago when my ligation reactions, once again, failed to produce viable colonies for me to expand. I wrapped things up in my notebook, noted things to do differently, and who knows, I might come back to it down the road.

Now that I'm an official member of the lab, I wanted to really dive into my ideas for thesis projects. I am especially interested in integration, which is necessary (but insufficient) for cancer progression. I am especially interested in the mechanisms underlying integration, which is not known. However, Jian's lab doesn't specialize or focus on these questions directly, so I wanted to take some time to read up on the relevant literature, define my questions and develop experimental approaches. Basically, come up with a thesis project.

So I've been reading non-stop for two weeks now, and sometimes I feel like my head is going to burst. It's kind of interesting to see how, over the course of a week, I have gotten to the point where I can pretty much breeze through the introduction (been there, done that) and go straight to the results and discussion. Sometimes I also spend a lot of time in the materials and methods section. It's been simultaneously super exciting and super boring. I am not doing any real lab work, which is boring, but I am learning a lot, making connections, developing ideas, and asking questions, which is very exciting.

I spent almost an hour and a half with Jian today talking about some of the papers I've been reading and refocusing my inquiries. We also started talking about experimental approaches which was useful but a little premature. I am hoping that after another week or so of reading I will be able to really home in on the questions I want to ask and then start coming up with ways of addressing them.

Additionally, I am giving lab meeting this week, which will be a summary of my rotation experience and a small taste of what's to come. This will probably mean I will get a little less reading done this week, but that's okay. I'm not exactly pressed for time or anything yet.

So far it's been a really great experience going through the literature, but I have quickly found that the paper trail never ends...almost every paper references another paper that I think is worth reading, which then references another paper. Sometimes you can pick out a few key papers in the field which are commonly referenced but I could literally spend a year reading up on everything HPV related and never get through it all. It's been hard trying to establish limits but I am getting better at it.

I will hopefully have more interesting things to share in the coming week(s) so stay tuned!

Grad Diary 3/22/10

For the past three weeks or so I have been unsuccessfully trying to insert a particular sequence into a plasmid vector containing the HPV16 genome. What makes this particular cloning experiment hard is that the vector is rather large (~10kb!) and my insert is not only small, but it is a mixture of various polymers of the same sequence (anywhere from five to over a hundred repeats). I don't know how many agar plates I went through (maybe twenty?) while I've been trying to get things to work. Jian has been helping me troubleshoot throughout the way to optimize the ligation reaction, DNA extraction methods (for example, is it better to do a gel extraction or the tried and true phenol:chloroform:isoamyl alochol extraction with ethanol precipitation?) It got to the point where I am literally running out of both vector and insert.

Finally, after working on this for weeks, I came in this morning and checked my plates and was surprised to see an explosion of growth! Unfortunately, even my ligation control plate (which is vector without insert) also showed an explosion of growth, so that means that one of the key steps that has been hampering my progress is the de-phosphorylation step with CIP. We ordered new CIP. Fortunately, this is still pretty good since, at first blush, it seems like my ligation reactions still had more growth than the controls. I re-plated my reactions at a much smaller volume, and hopefully they will have reasonably sized colonies for me tomorrow.

It was one of those moments where, after weeks of banging my head against the wall, there was suddenly an "Aha!" moment, and things looked like they are finally falling in place. There is still some work to do to see if there is anything usable, but even if I have to start over, I now have everything in place that it will only take a week to go from start to finish, instead of almost a month. Still, Jian and I have decided that I will stop after this week so I can focus more on reading and preparing my ideas for my thesis project(s). Maybe I can get one of the undergrads to finish things for me. It's kind of nice being able to say that, ha ha.

Thesis Lab Decision

As I am coming to the end of my third and final rotation in Jianxin's lab, I have come to the point where I have to decide which lab I would like to formally join as a full-time grad student.

Initially I thought this decision would be pretty easy; at the conclusion of my rotation with Una last semester, I was 99.999% sure I would join that lab, and I came into Jian's lab with the feeling that I would just enjoy myself and learn as much as I could.

As time progressed in Jian's lab, however, I became really fascinated by the HPV field and some of the unanswered questions, especially mechanisms underlying integration, a critical (and necessary) step for the onset of cervical cancer caused by the virus. And although my rotation project had nothing to do with this, I took time to read more and more and become even more excited about the possibilities. I also found that I was much more actively engaged during lab meetings, offering critical observations about the data and offering my ideas. In short, I have been much more engaged scientifically in Jian's lab than I ever was in Una's lab.

This past week I met with both PI's individually to seriously discuss potential thesis projects, expectations for me as a grad student, things I was interested in or would like to see happen in the lab if I were to join. I finished the week by meeting with Bob Doms, the chair of the microbiology department, and talking to him about my decision and listening to his advice, concerns, and suggestions. It was definitely helpful, and validated my own feelings about my choices and where I thought I wanted to be.

In the end, I decided to join Jian's lab. It feels like the best decision in pretty much every parameter I can think of - funding, research interests, support, etc - but that has not made coming to this decision or moving forward with it easier. My time with Una was fantastic, and it was, by the far, the best environment of people I could ever have hoped for - I hope I will be half as lucky finding labs that felt so much like a family to me and that worked so well together. But in the end, I am here to do research, to investigate and learn, and I think the Jian's lab offers me the best opportunities for all of that.

There are still risks and unknowns in joining Jian's lab: I will be her first graduate student (largely because she is so new here), and that brings with it some uncharted territory for both of us. But this is also an incredible opportunity for me to have a huge impact in the development of the lab, in attracting new students to the lab, in directing where the research goes, etc. There is also the risk in joining a lab with a faculty who hasn't formally received tenure yet. Normally I would stay far, far away from such a lab, because there is the risk the faculty member won't receive tenure while you are in the middle of your graduate work, and generally there is a stronger push to produce papers and get work done in the process of securing tenure. However, in this case, Jian is very well on her way towards receiving tenure and she has had a great team of post-docs who have been helping to establish the lab. Bob (the chair of Microbiology I mentioned above) has no doubt that she will receive tenure, and he probably has the largest voice in that decision out of anyone, so I trust his judgment. I think in this case, the risks involved in joining a junior faculty's lab are much less than other labs I see here at Penn.

I am extremely excited to count myself as Jian's first graduate student, and will be taking the next month or so to really develop my ideas for the questions I am interested in pursuing. Fortunately, a large component of my interest will greatly benefit from Una's expertise in HIV integration, and I have already talked to her about this collaboration. She was extremely excited about it, and I am glad to say that I foresee continuing working with her and her talented crew well into the future.

Grad Diary 3/9/10

I'm two days into my "spring break" and so far, it's been nice.

As I mentioned in my previous post, I don't have classes this week (and the undergrads are gone! no long lines at the carts for lunch!). In addition to taking the time to focus on lab stuff, I decided that I would take this week to seriously weigh my options between Una and Jian's lab, and try to come to a decision by the end of the week. To that end I've set up meeting times with both PI's (on different days) to talk seriously about potential thesis projects (both what they are interested as well as my own ideas) as well as wrapping up other concerns/expectations that come with being a full time graduate student.

Bob Doms, the chair for the Microbiology Department, has been very helpful. He works very closely with the graduate students who work in his faculties' labs (mostly MVP students) and meets with them at least once a semester (if not more) to check in, talk about upcoming decisions, etc. A few weeks ago he and I chatted about my vacillation between the two labs, and was happy with the extent and seriousness I have been considering the decision up till now. He did offer some helpful perspective and commentary, and we decided I would check in again when I thought I had made a decision. I will meet with him on Friday to talk about what I feel is the best decision for me (after having met with both PI's) and then make a "formal" announcement to both groups.

To that end I've been making lists and thinking about what the realities might be in both labs, the pros and cons, the differences in lab demeanor and philosophies, the opportunities, and the unknowns (recall that Jian doesn't have any full time grad students...I would be her first). I feel i am coming to a decision already, but I definitely want to take the time to hash it out with both PI's first.

I'll wait until next week to post again with my formal lab decision and how things are moving forward. Wish me luck!