So, today I had a lab meeting with Dr. Fraser. It was my first formal meeting with him since I've started in his lab almost a month ago. Iryna was with me too. It went pretty well, although in the beginning I had my doubts. His opening question was, "pretend I'm one of your classmates and explain to me what you're doing." This took me completely by surprise because I was expecting to give him a status update, not a mini talk. While I knew the basics of what I was doing, I didn't feel prepared to talk about all of the biology involved in my project - I would need to review some material before that, so I was a little flummoxed. It was also weird trying to give this little talk to the mastermind behind the whole project to begin with.
After stammering my way over some of the introductory stuff I caught my groove once I started talking about my projects, and things went smoothly after that. Predictably, Dr. Fraser said I should give more details on the background, but he was impressed with my explanation. He had specifically given me no warning before hand to see how I would do...apparently he does this a lot, especially to newbies.
He probed me a bit during my explanations to make sure I understood not only what I was doing but why. This wasn't a big problem because I generally make a commitment to knowing why things are done the way they are done. It's very easy, especially in microbiology, to get sucked into running protocols and not understanding why each step is needed or what the underlying scientific principles are.
I did run into one small crisis when I was talking about bacterial transformation. This is a process where you introduce novel genes into bacteria, which are very good at picking up foreign DNA and incorporating them into their genomes. While this can be a pain when bacteria share antibiotic resistance, it also makes it easy for us to engineer them to make things we want, like human insulin. Anyway, I was explaining how I was constructing my target plasmid by isolating my desired insert using restriction endonucleases and ligating them into a vector plasmid. One important step is dephosphorylating the vector plasmid after you've cut it open. I kind of brushed over this and Dr. Fraser asked me why we would dephosphorylate the vector and not the insert. It had been a while since I had reviewed the actual science behind restriction endonucleases and plasmid construction, and I was stumped for a moment.
Before letting myself panic, I calmly started to reason out loud why this would be the case. "Well, let's see. You need a phosphorylated 3' end if you're going to add a nucleotide since the polymerase doesn't add one, and you also need one if you're going to do ligation..." and as I said this in my mind I saw an endonuclease cut up my vector, making a nice open space for my insert. On the opposing 3' ends were bright yellow phosphate groups (phosphates are always yellow in biochem texts). I then pictured a DNA ligase coming in and before my insert was able to move in place, it neatly sealed up my vector back to its initial state. This all took probably a second and suddenly I knew the answer.
"Oh, of course. It's to keep the vector from re-ligating and excluding my insert."
"Very good!"
I felt a mixture of pride (for thinking quickly on my feet and being able to reason it out) and stupidity (for not knowing this sooner). All in all, Dr. Fraser was happy with my progress and impressed with my level of understanding, which made me feel pretty good.
Lessons of the day: 1) Make sure you understand the science behind everything you do. 2) If posed a question you don't know the answer to immediately, calmly reason it out using what you already know.
Wednesday, July 15, 2009
Monday, July 6, 2009
Who's Who - Fraser Lab
So it's been a bit longer than I initially wanted before another update, but now things are starting to slow down a bit. Today I'm going to introduce you to everyone in my lab and in the process explain the "hierarchy" of lab personnel that you may or may not be familiar with.
The set up for my lab is pretty typical for most labs, with the exception that there aren't any grad students in it except for me.
Some Background
For those without undergraduate backgrounds in biology/biochemistry skip this section unless you want to be bored to tears. This is just to give some context for what everyone in the lab is doing.
As I mentioned in my previous post, the Fraser lab studies mechanisms of latency in Herpes simplex 1 (HSV-1, currently there is no vaccine or cure). I also mentioned that Herpes simplex, when infecting epithelial cells (like skin) produces an acute infection with active replication and virion production. When these virions infect neuronal cells, however, they become latent (dormant) and do not actively transcribe most of their genes. No matter the cell type, HSV-1 DNA immediately becomes associated with histones in the host nucleus, which is thought to be a host defense mechanism to silence viral genomic material. For whatever reason (this is still being hotly investigated), in epithelial cells the histones are modified to look like "euchromatin," i.e. active replication still occurs (this is how your own genes are transcribed - they often still have histone proteins associated but in a modified state to allow for transcription). In neurons, however, these crucial histone modifications do not happen. Instead, the HSV-1 DNA becomes tightly associated with histones to form "heterochromatin" (i.e. the transcriptionally silent form of chromatin), though it does produce one transcript in high quantity. This transcript is an RNA species termed the Latency Associated Transcript (LAT), which exists as an extremely stable 1.5kb intron-like structure. It is thought that the LAT facilitates in keeping HSV-1 DNA transcriptionally silent. The exact mechanisms by which latency is relaxed in response to neuronal stress (to give rise to recurrent outbreaks characteristic of herpes infection) is not yet well understood.
With all that said you will now be able to understand what most of us are doing in the lab. I will now go down the "command line" of the lab starting with the PI going down to undergrads.
The Primary Investigator (PI)
My PI's name is Dr. Nigel Fraser, and he is a well known expert on Herpes simplex. His lab has been investigating mechanisms of HSV-1 latency for some time now. He has a one year sabbatical coming up this fall, and this summer he is busy attending conferences and setting up his sabbatical work. I don't see him much; when he's here I usually run into him once or twice a day, say hello, and maybe tell him what I am up to for the day. This is how a lot of labs function, as the PI's primary role is to manage his/her research group, and more importantly apply for funding and present at conferences. At this point I don't meet with him regularly for an update on what I'm doing, most likely because he's been out of the office half of the time thus far. Still, I know he is keeping an eye on me in other ways, and every now and then we will chat for fifteen or twenty minutes or so about what's going on in the lab and how my rotation is going.
Senior Research Assistant
A senior researcher is someone who has a PhD and a lot of experience but instead of heading their own lab, are working for another PI. This might be because of a long time as a collaborator or a really good post doc experience, etc. The senior researcher is often the second highest up person in the lab as far as experience and direction is considered, as s/he is directly below the PI. Our senior researcher is Jaewook Oh. Jaewook is Korean, and he is working on mapping the regions of the HSV-1 DNA that are commonly associated with histone proteins (nucelosomes). Right now this is being investigated using microarray technology which will allow him to probe the entire HSV-1 genome using DNA probes derived from HSV nucelosomes (this is done by purifying HSV DNA and then treating with Micrococcal nuclease, an enzyme which degrades DNA not associated with Histones). He also does a lot of ChIP (Chromosome ImmunoPrecipitation) and is investigating the histone modfications common in latent HSV DNA. I don't talk with him very much. His English is good but sometimes communicating about science kind of difficult at times (he gave a lab meeting presentation last week which at times was kind of hard to follow for this reason). He seems like an overall nice guy but when he needs to do something you better get out of his way because he can be pretty direct at times.
The Post-Doc's
A "postdoc" is a "post-doctoral fellow" which, as the name implies, is someone who has completed their PhD training and is now working in a lab as a fellowship. These fellowships are generally for two years, and are a way to get very intensive training in a technique the lab is famous for, or for exploring a very specific model or question. When full professors go on sabbatical, they sometimes do something similar (usually for a year) where they spend time in a lab to hone their skills or get a new perspective. It is expected that researchers will have at least one postdoc experience before going into industry, and two or more if you want to go into academia. In research labs, the post docs are often the "lieutenants" of the lab, in charge of very specific projects, and they often have a grad student working with them. They are considered full time employees of the PI. Our lab has a rather large number of postdocs (4). They are:
Kerry - Kerry is investigating the stability and properties of the LAT intron species. The LAT transcript can be alternatively spliced into different forms with varying structures (the major species is a lariot common to spliced-out introns but is not degraded). Her work is more genetically oriented, which is pretty cool. I don't talk with her very much either, and like Jaewook she seems nice overall but can be a little rough around the edges when she's working or needs to get something done.
Gerald - Gerald just started in the Fraser lab a month before I came. He is actually investigating the uses of HSV as a possible gene vector for neuronal-related diseases (since HSV targets neurons so efficiently). His work is more neuroscience releated and he will be working a lot with mice. He is really easy to talk to and quick to laugh, and is really approachable if you have a question about something. His wife just gave birth to their second child (a girl) last Wednesday, so we are all pretty excited for him.
Ketna - Ketna just joined our lab last Wednesday. I have no idea what she will be working on, though she's been working with Gerald a lot these past few days. She seems nice, though I haven't had a chance to talk with her past my introduction. I'm hoping to catch lunch with her this week to have a longer chat. At this point her work is pretty boring as she will be doing a LOT of background reading (I'm still doing that now!).
Iryna - Iryna is originally from the Ukraine. She earned her PhD at the University of Oklahoma doing lots of genetics work, which means she has a similar background as I do (in that she has more experience with genetics and is learning virology). I have been assigned to Iryna for the summer, which thus far has worked out really well, as she also commutes into the city every day, and our schedules match up well. At first I thought she was kind of distant and direct, but then learned it was more of a function of English being her second language. She is actually fun to talk and joke with and has been a good match for me thus far in that she expects good work from me but let's me make mistakes too. Right now she is working on two projects; she is collaborating with Jaewook (above) to prepare the DNA probes for the microarray. Her major project is developing an HSV-1 strain that, when latenlty infected, can be visualized in an easy way (i.e. latently infected cells can be spotted under the microscope). Since latenly infected cells produce only LAT transcript (at high levels) this transcript is a great candidate as a molecular marker. Iryna is trying to introduce a construct with GFP (green fluorescent protein) sequence dowstream of the LAT promoter, which will create a GFP-LAT fusion protein. If this works, the virus will produce fluorescently labeled LAT's that can be visualized in a microscope (and sorted). She is currently tyring to do this via homologous recombination using HSV DNA and a plasmid with her LAT-GFP construct, which has a very low success rate (1%). If she can do it, though, it will be an extremely useful tool for studying latenly infected neurons.
Lab Technicians
The lab techs are sometimes the most neglected people in the lab, which is a shame because they are the people keeping the labs running smoothly, and they often have the most expertise with the lab's research questions (often more than the post docs). Lab techs are often people just out of undergrad or who have their master's. They do a lot of the grunt work for the lab like preparing solutions, ordering reagents, maintaining cell lines and growing virus. They often have side projects that they work on as well. They are an indispensable part of the team. Our lab tech's name is Jared. I thought he was not much older than me, and was surprised to learn he was 28. He's been working in the Fraser lab for almost seven years now, and he has a lot of expertise. He's also really fun and approachable. He went out of his way to make sure I was all set up and had everything I needed, and he has been very patient telling me how things are run in the lab (how do I dispose of this? where can I get more pipette tips? Where do you store ethanol?) He's very good at what he does, is very efficient and organized, but also the main source of lab hilarity. He's somewhat of a joker and keeps the lab environment light. He also commutes into the city and sometimes we walk back to 30th street station together after work. I feel really comfortable going to him for help (often!) and it's great having someone in the lab that you can turn to without worrying about stepping on someone's toes.
We also have a lab tech in training, Sindura, who is from India. If I remember correctly she got her Masters in biomedical engineering and is currently working with Jaewook on his project and with Jared to learn how to be a technician. She's nice to talk to, and someone else I can turn to if I have a question about where something is in the lab, though when it comes to questions regarding viral preps or protocols I still turn to Jared.
Grad Student(s)
Generally there will be a grad student or two working on their thesis project(s). Dr. Fraser doesn't currently have any grad students doing thesis work, just me doing my summer rotation which is a temporary affair anyway. You already know about me, but as far as what I'm actually doing: I am mainly learning as many techniques as possible in the lab, including growing and maintaining cells in culture, growing up virus, titering virus (purifying and quantifying virus), producing recombinant virus, etc. As far as my project is concerned, I will basically be following Iryna's work in parallel, except instead of using GFP I will be making a fusion protein with a related fluorescent protein called "mCherry." The disadvantage with using GFP is that when cells die they sometimes autofluoresce in a wavelength similar to GFP, so you can get false positives. Using mCherry (which is, predictably, red) eliminates this. We'll see how far I can get.
Undergrad(s)
Sometimes you have undergrads in the lab working on little projects to get their feet wet. At a large research university like Penn, it is usually not nearly as intensive as my experience was at Ithaca because the labs are so much bigger. Right now there are two undergrads doing a work study. There is Prioty, who was born in Bangladesh but lived most of her life in Canada. She's an undergrad at Penn, and she's studying for the MCAT's right now (God bless her) and is pretty smart. Prioty is working with Kerry on the LAT transcript. There is also Jenny, who is doing a workstudy through Drexel (which is famous for their co-ops) and is working closely with Jared. Jenny is from Vietnam originally. She's really nice and a little timid around the lab, probably because she feels everyone around her is much more experienced than her. I try to keep her at ease as much as possible. It's kind of strange to already be seeing undergrads and not count myself one of them, but I can honestly say the transition has been pretty rapid. Anyway, Prioty and Jenny are good workers and fun to talk with, and it's nice to know I'm not the only one who is trying to figure out what the hell is going on.
And that is pretty much everyone! It's quite a group! I feel fortunate that there aren't any ridiculous personality issues within the lab, which can be a real pain. Everyone pretty much keeps to themselves, though there is some soicializing every now and then (usually with the undergrads, Jared and/or Gerald). I really like the people I'm working with and feel that, while I have a lot to learn, I have the resources and the environment I need to do well and enjoy myself.
Looking forward to sharing more in the coming days. Hope you all had a great fourth of July and I hope to hear from you soon!
The set up for my lab is pretty typical for most labs, with the exception that there aren't any grad students in it except for me.
Some Background
For those without undergraduate backgrounds in biology/biochemistry skip this section unless you want to be bored to tears. This is just to give some context for what everyone in the lab is doing.
As I mentioned in my previous post, the Fraser lab studies mechanisms of latency in Herpes simplex 1 (HSV-1, currently there is no vaccine or cure). I also mentioned that Herpes simplex, when infecting epithelial cells (like skin) produces an acute infection with active replication and virion production. When these virions infect neuronal cells, however, they become latent (dormant) and do not actively transcribe most of their genes. No matter the cell type, HSV-1 DNA immediately becomes associated with histones in the host nucleus, which is thought to be a host defense mechanism to silence viral genomic material. For whatever reason (this is still being hotly investigated), in epithelial cells the histones are modified to look like "euchromatin," i.e. active replication still occurs (this is how your own genes are transcribed - they often still have histone proteins associated but in a modified state to allow for transcription). In neurons, however, these crucial histone modifications do not happen. Instead, the HSV-1 DNA becomes tightly associated with histones to form "heterochromatin" (i.e. the transcriptionally silent form of chromatin), though it does produce one transcript in high quantity. This transcript is an RNA species termed the Latency Associated Transcript (LAT), which exists as an extremely stable 1.5kb intron-like structure. It is thought that the LAT facilitates in keeping HSV-1 DNA transcriptionally silent. The exact mechanisms by which latency is relaxed in response to neuronal stress (to give rise to recurrent outbreaks characteristic of herpes infection) is not yet well understood.
With all that said you will now be able to understand what most of us are doing in the lab. I will now go down the "command line" of the lab starting with the PI going down to undergrads.
The Primary Investigator (PI)
My PI's name is Dr. Nigel Fraser, and he is a well known expert on Herpes simplex. His lab has been investigating mechanisms of HSV-1 latency for some time now. He has a one year sabbatical coming up this fall, and this summer he is busy attending conferences and setting up his sabbatical work. I don't see him much; when he's here I usually run into him once or twice a day, say hello, and maybe tell him what I am up to for the day. This is how a lot of labs function, as the PI's primary role is to manage his/her research group, and more importantly apply for funding and present at conferences. At this point I don't meet with him regularly for an update on what I'm doing, most likely because he's been out of the office half of the time thus far. Still, I know he is keeping an eye on me in other ways, and every now and then we will chat for fifteen or twenty minutes or so about what's going on in the lab and how my rotation is going.
Senior Research Assistant
A senior researcher is someone who has a PhD and a lot of experience but instead of heading their own lab, are working for another PI. This might be because of a long time as a collaborator or a really good post doc experience, etc. The senior researcher is often the second highest up person in the lab as far as experience and direction is considered, as s/he is directly below the PI. Our senior researcher is Jaewook Oh. Jaewook is Korean, and he is working on mapping the regions of the HSV-1 DNA that are commonly associated with histone proteins (nucelosomes). Right now this is being investigated using microarray technology which will allow him to probe the entire HSV-1 genome using DNA probes derived from HSV nucelosomes (this is done by purifying HSV DNA and then treating with Micrococcal nuclease, an enzyme which degrades DNA not associated with Histones). He also does a lot of ChIP (Chromosome ImmunoPrecipitation) and is investigating the histone modfications common in latent HSV DNA. I don't talk with him very much. His English is good but sometimes communicating about science kind of difficult at times (he gave a lab meeting presentation last week which at times was kind of hard to follow for this reason). He seems like an overall nice guy but when he needs to do something you better get out of his way because he can be pretty direct at times.
The Post-Doc's
A "postdoc" is a "post-doctoral fellow" which, as the name implies, is someone who has completed their PhD training and is now working in a lab as a fellowship. These fellowships are generally for two years, and are a way to get very intensive training in a technique the lab is famous for, or for exploring a very specific model or question. When full professors go on sabbatical, they sometimes do something similar (usually for a year) where they spend time in a lab to hone their skills or get a new perspective. It is expected that researchers will have at least one postdoc experience before going into industry, and two or more if you want to go into academia. In research labs, the post docs are often the "lieutenants" of the lab, in charge of very specific projects, and they often have a grad student working with them. They are considered full time employees of the PI. Our lab has a rather large number of postdocs (4). They are:
Kerry - Kerry is investigating the stability and properties of the LAT intron species. The LAT transcript can be alternatively spliced into different forms with varying structures (the major species is a lariot common to spliced-out introns but is not degraded). Her work is more genetically oriented, which is pretty cool. I don't talk with her very much either, and like Jaewook she seems nice overall but can be a little rough around the edges when she's working or needs to get something done.
Gerald - Gerald just started in the Fraser lab a month before I came. He is actually investigating the uses of HSV as a possible gene vector for neuronal-related diseases (since HSV targets neurons so efficiently). His work is more neuroscience releated and he will be working a lot with mice. He is really easy to talk to and quick to laugh, and is really approachable if you have a question about something. His wife just gave birth to their second child (a girl) last Wednesday, so we are all pretty excited for him.
Ketna - Ketna just joined our lab last Wednesday. I have no idea what she will be working on, though she's been working with Gerald a lot these past few days. She seems nice, though I haven't had a chance to talk with her past my introduction. I'm hoping to catch lunch with her this week to have a longer chat. At this point her work is pretty boring as she will be doing a LOT of background reading (I'm still doing that now!).
Iryna - Iryna is originally from the Ukraine. She earned her PhD at the University of Oklahoma doing lots of genetics work, which means she has a similar background as I do (in that she has more experience with genetics and is learning virology). I have been assigned to Iryna for the summer, which thus far has worked out really well, as she also commutes into the city every day, and our schedules match up well. At first I thought she was kind of distant and direct, but then learned it was more of a function of English being her second language. She is actually fun to talk and joke with and has been a good match for me thus far in that she expects good work from me but let's me make mistakes too. Right now she is working on two projects; she is collaborating with Jaewook (above) to prepare the DNA probes for the microarray. Her major project is developing an HSV-1 strain that, when latenlty infected, can be visualized in an easy way (i.e. latently infected cells can be spotted under the microscope). Since latenly infected cells produce only LAT transcript (at high levels) this transcript is a great candidate as a molecular marker. Iryna is trying to introduce a construct with GFP (green fluorescent protein) sequence dowstream of the LAT promoter, which will create a GFP-LAT fusion protein. If this works, the virus will produce fluorescently labeled LAT's that can be visualized in a microscope (and sorted). She is currently tyring to do this via homologous recombination using HSV DNA and a plasmid with her LAT-GFP construct, which has a very low success rate (1%). If she can do it, though, it will be an extremely useful tool for studying latenly infected neurons.
Lab Technicians
The lab techs are sometimes the most neglected people in the lab, which is a shame because they are the people keeping the labs running smoothly, and they often have the most expertise with the lab's research questions (often more than the post docs). Lab techs are often people just out of undergrad or who have their master's. They do a lot of the grunt work for the lab like preparing solutions, ordering reagents, maintaining cell lines and growing virus. They often have side projects that they work on as well. They are an indispensable part of the team. Our lab tech's name is Jared. I thought he was not much older than me, and was surprised to learn he was 28. He's been working in the Fraser lab for almost seven years now, and he has a lot of expertise. He's also really fun and approachable. He went out of his way to make sure I was all set up and had everything I needed, and he has been very patient telling me how things are run in the lab (how do I dispose of this? where can I get more pipette tips? Where do you store ethanol?) He's very good at what he does, is very efficient and organized, but also the main source of lab hilarity. He's somewhat of a joker and keeps the lab environment light. He also commutes into the city and sometimes we walk back to 30th street station together after work. I feel really comfortable going to him for help (often!) and it's great having someone in the lab that you can turn to without worrying about stepping on someone's toes.
We also have a lab tech in training, Sindura, who is from India. If I remember correctly she got her Masters in biomedical engineering and is currently working with Jaewook on his project and with Jared to learn how to be a technician. She's nice to talk to, and someone else I can turn to if I have a question about where something is in the lab, though when it comes to questions regarding viral preps or protocols I still turn to Jared.
Grad Student(s)
Generally there will be a grad student or two working on their thesis project(s). Dr. Fraser doesn't currently have any grad students doing thesis work, just me doing my summer rotation which is a temporary affair anyway. You already know about me, but as far as what I'm actually doing: I am mainly learning as many techniques as possible in the lab, including growing and maintaining cells in culture, growing up virus, titering virus (purifying and quantifying virus), producing recombinant virus, etc. As far as my project is concerned, I will basically be following Iryna's work in parallel, except instead of using GFP I will be making a fusion protein with a related fluorescent protein called "mCherry." The disadvantage with using GFP is that when cells die they sometimes autofluoresce in a wavelength similar to GFP, so you can get false positives. Using mCherry (which is, predictably, red) eliminates this. We'll see how far I can get.
Undergrad(s)
Sometimes you have undergrads in the lab working on little projects to get their feet wet. At a large research university like Penn, it is usually not nearly as intensive as my experience was at Ithaca because the labs are so much bigger. Right now there are two undergrads doing a work study. There is Prioty, who was born in Bangladesh but lived most of her life in Canada. She's an undergrad at Penn, and she's studying for the MCAT's right now (God bless her) and is pretty smart. Prioty is working with Kerry on the LAT transcript. There is also Jenny, who is doing a workstudy through Drexel (which is famous for their co-ops) and is working closely with Jared. Jenny is from Vietnam originally. She's really nice and a little timid around the lab, probably because she feels everyone around her is much more experienced than her. I try to keep her at ease as much as possible. It's kind of strange to already be seeing undergrads and not count myself one of them, but I can honestly say the transition has been pretty rapid. Anyway, Prioty and Jenny are good workers and fun to talk with, and it's nice to know I'm not the only one who is trying to figure out what the hell is going on.
And that is pretty much everyone! It's quite a group! I feel fortunate that there aren't any ridiculous personality issues within the lab, which can be a real pain. Everyone pretty much keeps to themselves, though there is some soicializing every now and then (usually with the undergrads, Jared and/or Gerald). I really like the people I'm working with and feel that, while I have a lot to learn, I have the resources and the environment I need to do well and enjoy myself.
Looking forward to sharing more in the coming days. Hope you all had a great fourth of July and I hope to hear from you soon!
Wednesday, June 24, 2009
Grad Diary 6/24/09
This afternoon I found myself in a rather delicate when Prioty, an undergrad who is doing a workstudy in our lab this summer, asked me a seemingly innocuous question: what voltage do I normally run my agarose gels?
True, it seems like a simple enough question (unless you have no idea what electrophoresis is), unless you knew why she was asking me. You see, Prioty is an undergrad and is working under one of the post-docs in the lab, Kerry. A post doc is someone who already has their PhD but is getting specialized training in a lab, usually for two or three years. Anyway, Prioty has been working with Kerry for the past few months, and recently she's been running agarose gels of radiolabeled DNA. Things had been going smoothly, but recently her gels were showing poor results - there was streaking, the bands were bowed like smiley faces, or sometimes she wouldn't get bands at all. For the fifteen minutes before Prioty asked me for my input, I could easily overhear the two of them discussing her latest gel, and Kerry was trying to diagnose where the problem step was (the buffer? the voltage? pouring the gel?). Kerry is pretty good at what she does, but she is not a very good teacher, and she isn't very patient...she knows how she does it but doesn't do a good job of explaining why she does it that way.
So after having this conversation with Kerry, Prioty was told that she should run the gel at a lower voltage.
I could easily overhear all of this going on (they are right behind my bench) so I knew exactly why Prioty asked me what voltage I normally run my gels. On the one hand she wanted to get help, but really she wanted to get back at Kerry by getting my approval for how she was running the gel.
I found this position to be kind of amusing - I mean, she obviously thought I was smart enough to be a credible source of advice - but at the other hand I didn't appreciate the passive aggressive route she was taking in her relationship with Kerry. So I tried to be as helpful as I could without giving Prioty any ammo to shoot at Kerry (the last thing I needed was to get on anyone's bad side, I've only been here a few weeks!).
Fortunately, when it comes to doing electrophoresis, the voltage depends on a lot of factors: the size of the fragments you're interested in, the concentration of the gel you've poured, the buffer you're using, how well you need to separate bands of differing sizes. So I was able to tell Prioty quite truthfully that, though she should take all these things into account, the best person to ask would be someone who actually has experience with her particular experiment - i.e. Kerry.
While I obviously didn't give her what she really wanted, she seemed satisfied with this answer and went on her way. All in all, I found the entire thing quite amusing, though I also learned a valuable lesson; it's sometimes important to take note of the dynamics of the lab and not always be caught up with what you are doing.
One word to those just getting into research; if you disagree with someone about how best to do an experiment, be open about it. It's much better to find out why someone does an experiment a particular way and to talk about the relative merits of doing it differently rather than try to prove you're smarter than the other person.
Cheers!
True, it seems like a simple enough question (unless you have no idea what electrophoresis is), unless you knew why she was asking me. You see, Prioty is an undergrad and is working under one of the post-docs in the lab, Kerry. A post doc is someone who already has their PhD but is getting specialized training in a lab, usually for two or three years. Anyway, Prioty has been working with Kerry for the past few months, and recently she's been running agarose gels of radiolabeled DNA. Things had been going smoothly, but recently her gels were showing poor results - there was streaking, the bands were bowed like smiley faces, or sometimes she wouldn't get bands at all. For the fifteen minutes before Prioty asked me for my input, I could easily overhear the two of them discussing her latest gel, and Kerry was trying to diagnose where the problem step was (the buffer? the voltage? pouring the gel?). Kerry is pretty good at what she does, but she is not a very good teacher, and she isn't very patient...she knows how she does it but doesn't do a good job of explaining why she does it that way.
So after having this conversation with Kerry, Prioty was told that she should run the gel at a lower voltage.
I could easily overhear all of this going on (they are right behind my bench) so I knew exactly why Prioty asked me what voltage I normally run my gels. On the one hand she wanted to get help, but really she wanted to get back at Kerry by getting my approval for how she was running the gel.
I found this position to be kind of amusing - I mean, she obviously thought I was smart enough to be a credible source of advice - but at the other hand I didn't appreciate the passive aggressive route she was taking in her relationship with Kerry. So I tried to be as helpful as I could without giving Prioty any ammo to shoot at Kerry (the last thing I needed was to get on anyone's bad side, I've only been here a few weeks!).
Fortunately, when it comes to doing electrophoresis, the voltage depends on a lot of factors: the size of the fragments you're interested in, the concentration of the gel you've poured, the buffer you're using, how well you need to separate bands of differing sizes. So I was able to tell Prioty quite truthfully that, though she should take all these things into account, the best person to ask would be someone who actually has experience with her particular experiment - i.e. Kerry.
While I obviously didn't give her what she really wanted, she seemed satisfied with this answer and went on her way. All in all, I found the entire thing quite amusing, though I also learned a valuable lesson; it's sometimes important to take note of the dynamics of the lab and not always be caught up with what you are doing.
One word to those just getting into research; if you disagree with someone about how best to do an experiment, be open about it. It's much better to find out why someone does an experiment a particular way and to talk about the relative merits of doing it differently rather than try to prove you're smarter than the other person.
Cheers!
Sunday, June 21, 2009
First Week of Summer Rotation
And so it begins!
My years as an undergraduate are now over (as of May 17), and graduation was followed by about four weeks of relaxation, visiting friends, having a fantastic graduation party, and gearing up for my summer rotation, which started this past week (Monday June 15).
Most everyone I spoke to had advised me to take the summer off to relax and enjoy myself before I entered the "real world." Granted, some people may not think that graduate school is the "real world" (and to an extent they are right), but it very much is the real world in the sense that I am entering into a lifestyle that will demand 50+ hours a week of work, odd work hours, little vacation time, and for the summer at least, a long commute. Anyway, I don't have a lot of money to be traveling this summer and most of my friends are getting jobs now, so the prospect of sitting at home all summer and working a part time job did not seem appealing. Since my efforts to secure a summer internship didn't pan out, I decided I would do an early rotation during the summer.
During the first year of grad school we will rotate through three different labs in an effort to find a lab/research advisor/project that we want to continue for our thesis. These are ten week appointments where we learn the techniques being used in the lab, get a feel for the project(s) going on there and the way the people in that lab work together (or don't) and even get a small project of our own. Because these rotations generally happen while we also have classes, we aren't expected to produce results or get a lot of great data; rather we are expected to get a feel as to whether this lab works well for us or not.
We have the option to do one of these rotations during the summer. I decided to take advantage of this for a variety of reasons: a) I will be paid for it (as I will be in general as a grad student), b) I will get a rotation out of the way, leaving me with more flexible time during the upcoming academic year, c) this rotation will not be burdneded with class time, giving me the whole day to learn techniques and work on projects, and d) my transition to Philadelphia and Penn will be earlier and smoother once classes start in the fall.
Since I live close enough to Philly to commute, I decided to hold off on getting an apartment until sometime during the summer. I am currently looking at apartments and am looking at moving in early August. This means that for the time being I am commuting to Penn, which thus far has been manageable, if exhausting. I take the R5 train into Philadelphia, which takes about 45 minutes from Ambler Station (a twenty five minute drive from home), and then walk from 30th St Station to my lab, a fifteen minute walk mostly through Penn's beautiful campus, making my total commute from my house to my lab approximately an hour and a half. Half of that is riding the train though, which is a great break and an opportunity to read, listen to music/podcasts, and just prepare myself for the day or begin unwinding. I'll be honest though, it's a LONG day, about twelve ours including commute time, but I'm slowly adjusting. Needless to say, I am looking forward to moving into the city soon, though living at home does have its perks ;)
I am working this summer in Dr. Nigel Fraser's lab. He is a molecular virologist (exactly what I want to do) and he works with Herpes simplex-1 (Herpes simplex-2 is the sexually transmitted one very similar to HSV-1). In layman's terms, one of the things he is looking into is the underlying mechanisms by which HSV becomes latent (dormant) in certain host cells. This latency strategy is what makes Herpes simplex a chronic disease that periodically "comes back."
For the more scientifically inclined:
HSV-1 establishes an acute infection when it encounters epithelial cells, producing blistering/cold sores commonly assoicated with the disease. During this acute infection, the active production of new virions eventually causes the infection of nearby neurons. When infecting neurons, a latent infection is established where lytic phase genes are repressed. This latent infection can be reactivated by means that are still under investigation. What makes Herpes very interesting is that when it's genomic material enters the host cell nucelus (Herpes is a DNA virus), the host cell will wrap up the HSV DNA in histones and chromatin in an effort to silence the foreign DNA. The Herpes virus carries with it proteins that then modify this chromatin formation to either relax histone binding for an acute infection in epithelial cells, or which supplement histone association to form a latent infection in neurons. We are still trying to figure out all the proteins involved in the decision to go into lytic or latent mode, and to maintain these decisions. Additionally, in latently infected cells there exists a Latency Associated Transcript (LAT), an RNA species that helps to maintain latency by mechanisms still under investigation. It's pretty amazing how HSV has developed a response to histone association, which would normally completely shut off HSV DNA and inhibit viral propagation. I love it.
Anyway, the lab uses a lot of basic virology (growing up host cells in tissue culture, infecting with virus, manipulating viral DNA, growing up and titering virus, etc) which will give me a good foundation in viral methodology. And it also does a lot of DNA manipulation that I am already pretty familiar with after working for so long in a genetics lab. I am working closely with a post-doc in the lab, Dr. Iryna Sanders, who is working on two projects: mapping the regions of HSV DNA that are closely associated with histones, and generating a genetically engineered virus that will express a GFP-tagged LAT, an RNA transcript associated with the maintenance of latency in neurons (see above). I will be helping her with these projects, and my own project will be a parallel of the GFP tagging project, but using a different fluorescent tag (m-Cherry).
So, this past week was my introduction into the Fraser lab. I technically started on Tuesday because on Monday everyone was at a huge Herpes virus conference held on the Penn campus while I went to summer orientation and took care of other errands. I am working very little with Dr. Fraser directly; rather I have been assigned to Dr. Iryna Sanders, one of three post-docs in the Fraser lab. She earned her PhD last spring and just started her virology training with Dr. Fraser last year, and so she's still learning too (though obviously quite a few steps ahead of me). She also has a relatively long commute, so our schedules will mesh quite well.
This week has been kind of crazy. I have a desk to myself, though I don't really have my own bench quite yet (I will probably get my space cleared next week). This whole week I've been shadowing Iryna in her virology work, trying to strike a balance of asking questions and observing, and then furiously taking notes. Fortunately I have enough lab experience to have a sense of what I should just watch and internalize (for example, certain tricks in tissue culture) and what to write down (protocol steps, centrifugation conditions, where things are stored, etc). I've also been given plenty of background reading material to get me up to speed, which has been a bear to get through, but very interesting. It's also kind of hard to keep track of everything since Iryna is working on two different projects simultaneously, but by the end of the week I've come up with a system to organize all my notes and my notebook. I'm also getting a hold of as many protocols as I can, and typing up my own (especially after editing the written protocol with what Iryna actually does). It's been a little overwhelming at times but all so exciting.
This week was also a test of whether my education at Ithaca has adequately prepared me for "the real deal," and I can honestly say that I am more than prepared for my work here, even though my experience with virology is limited. When Iryna was explaining the background and her projects, I was pleasantly suprised to find that I was able to follow all of the basic science and the rationale for the experiments quite easily, while small particulars (like the names of cell lines, certain viral proteins and other more specialized terms) were still unfamiliar but easily picked up after some background reading. It was very empowering to hear things like "we are currently working up DNA for MNase digestion so we can use a microarray to map the histone association sites" and not need an explanation of what MNase or microarrays are or why one would use those techniques (even though I've never personally done those experiments). Even the papers I've been reading as background haven't been too bad to get through, though they are mostly reviews and not too experimentally heavy.
There are two other post-docs in the lab with their own students, but they pretty much keep to themselves working on their own projects. I've introduced myself to all of them briefly but haven't really gotten to know them well yet. The lab technician, Jared, has been very helpful and friendly thus far, checking in daily to make sure I have everything I need. He and Iryna are the two people I talk to the most. For the most part everyone does their own thing, though they are also relatively approachable. I will probably talk more about their personalities in a few weeks when I have a better idea of who they are. All in all, it's pretty comfortable in the lab, if also a bit individualistic. I see Dr. Fraser in passing most days. I will be sitting in my first lab meeting next Tuesday, which should prove to be interesting.
I think that should do it for today. In the coming days I'll talk more about some of the social events I've attended so far, getting to know the other grad students in my program(s) and stuff about Penn and Philly. I'm looking forward to what the summer will bring both in the lab and out. Hope you all are enjoying your summer!
My years as an undergraduate are now over (as of May 17), and graduation was followed by about four weeks of relaxation, visiting friends, having a fantastic graduation party, and gearing up for my summer rotation, which started this past week (Monday June 15).
Most everyone I spoke to had advised me to take the summer off to relax and enjoy myself before I entered the "real world." Granted, some people may not think that graduate school is the "real world" (and to an extent they are right), but it very much is the real world in the sense that I am entering into a lifestyle that will demand 50+ hours a week of work, odd work hours, little vacation time, and for the summer at least, a long commute. Anyway, I don't have a lot of money to be traveling this summer and most of my friends are getting jobs now, so the prospect of sitting at home all summer and working a part time job did not seem appealing. Since my efforts to secure a summer internship didn't pan out, I decided I would do an early rotation during the summer.
During the first year of grad school we will rotate through three different labs in an effort to find a lab/research advisor/project that we want to continue for our thesis. These are ten week appointments where we learn the techniques being used in the lab, get a feel for the project(s) going on there and the way the people in that lab work together (or don't) and even get a small project of our own. Because these rotations generally happen while we also have classes, we aren't expected to produce results or get a lot of great data; rather we are expected to get a feel as to whether this lab works well for us or not.
We have the option to do one of these rotations during the summer. I decided to take advantage of this for a variety of reasons: a) I will be paid for it (as I will be in general as a grad student), b) I will get a rotation out of the way, leaving me with more flexible time during the upcoming academic year, c) this rotation will not be burdneded with class time, giving me the whole day to learn techniques and work on projects, and d) my transition to Philadelphia and Penn will be earlier and smoother once classes start in the fall.
Since I live close enough to Philly to commute, I decided to hold off on getting an apartment until sometime during the summer. I am currently looking at apartments and am looking at moving in early August. This means that for the time being I am commuting to Penn, which thus far has been manageable, if exhausting. I take the R5 train into Philadelphia, which takes about 45 minutes from Ambler Station (a twenty five minute drive from home), and then walk from 30th St Station to my lab, a fifteen minute walk mostly through Penn's beautiful campus, making my total commute from my house to my lab approximately an hour and a half. Half of that is riding the train though, which is a great break and an opportunity to read, listen to music/podcasts, and just prepare myself for the day or begin unwinding. I'll be honest though, it's a LONG day, about twelve ours including commute time, but I'm slowly adjusting. Needless to say, I am looking forward to moving into the city soon, though living at home does have its perks ;)
I am working this summer in Dr. Nigel Fraser's lab. He is a molecular virologist (exactly what I want to do) and he works with Herpes simplex-1 (Herpes simplex-2 is the sexually transmitted one very similar to HSV-1). In layman's terms, one of the things he is looking into is the underlying mechanisms by which HSV becomes latent (dormant) in certain host cells. This latency strategy is what makes Herpes simplex a chronic disease that periodically "comes back."
For the more scientifically inclined:
HSV-1 establishes an acute infection when it encounters epithelial cells, producing blistering/cold sores commonly assoicated with the disease. During this acute infection, the active production of new virions eventually causes the infection of nearby neurons. When infecting neurons, a latent infection is established where lytic phase genes are repressed. This latent infection can be reactivated by means that are still under investigation. What makes Herpes very interesting is that when it's genomic material enters the host cell nucelus (Herpes is a DNA virus), the host cell will wrap up the HSV DNA in histones and chromatin in an effort to silence the foreign DNA. The Herpes virus carries with it proteins that then modify this chromatin formation to either relax histone binding for an acute infection in epithelial cells, or which supplement histone association to form a latent infection in neurons. We are still trying to figure out all the proteins involved in the decision to go into lytic or latent mode, and to maintain these decisions. Additionally, in latently infected cells there exists a Latency Associated Transcript (LAT), an RNA species that helps to maintain latency by mechanisms still under investigation. It's pretty amazing how HSV has developed a response to histone association, which would normally completely shut off HSV DNA and inhibit viral propagation. I love it.
Anyway, the lab uses a lot of basic virology (growing up host cells in tissue culture, infecting with virus, manipulating viral DNA, growing up and titering virus, etc) which will give me a good foundation in viral methodology. And it also does a lot of DNA manipulation that I am already pretty familiar with after working for so long in a genetics lab. I am working closely with a post-doc in the lab, Dr. Iryna Sanders, who is working on two projects: mapping the regions of HSV DNA that are closely associated with histones, and generating a genetically engineered virus that will express a GFP-tagged LAT, an RNA transcript associated with the maintenance of latency in neurons (see above). I will be helping her with these projects, and my own project will be a parallel of the GFP tagging project, but using a different fluorescent tag (m-Cherry).
So, this past week was my introduction into the Fraser lab. I technically started on Tuesday because on Monday everyone was at a huge Herpes virus conference held on the Penn campus while I went to summer orientation and took care of other errands. I am working very little with Dr. Fraser directly; rather I have been assigned to Dr. Iryna Sanders, one of three post-docs in the Fraser lab. She earned her PhD last spring and just started her virology training with Dr. Fraser last year, and so she's still learning too (though obviously quite a few steps ahead of me). She also has a relatively long commute, so our schedules will mesh quite well.
This week has been kind of crazy. I have a desk to myself, though I don't really have my own bench quite yet (I will probably get my space cleared next week). This whole week I've been shadowing Iryna in her virology work, trying to strike a balance of asking questions and observing, and then furiously taking notes. Fortunately I have enough lab experience to have a sense of what I should just watch and internalize (for example, certain tricks in tissue culture) and what to write down (protocol steps, centrifugation conditions, where things are stored, etc). I've also been given plenty of background reading material to get me up to speed, which has been a bear to get through, but very interesting. It's also kind of hard to keep track of everything since Iryna is working on two different projects simultaneously, but by the end of the week I've come up with a system to organize all my notes and my notebook. I'm also getting a hold of as many protocols as I can, and typing up my own (especially after editing the written protocol with what Iryna actually does). It's been a little overwhelming at times but all so exciting.
This week was also a test of whether my education at Ithaca has adequately prepared me for "the real deal," and I can honestly say that I am more than prepared for my work here, even though my experience with virology is limited. When Iryna was explaining the background and her projects, I was pleasantly suprised to find that I was able to follow all of the basic science and the rationale for the experiments quite easily, while small particulars (like the names of cell lines, certain viral proteins and other more specialized terms) were still unfamiliar but easily picked up after some background reading. It was very empowering to hear things like "we are currently working up DNA for MNase digestion so we can use a microarray to map the histone association sites" and not need an explanation of what MNase or microarrays are or why one would use those techniques (even though I've never personally done those experiments). Even the papers I've been reading as background haven't been too bad to get through, though they are mostly reviews and not too experimentally heavy.
There are two other post-docs in the lab with their own students, but they pretty much keep to themselves working on their own projects. I've introduced myself to all of them briefly but haven't really gotten to know them well yet. The lab technician, Jared, has been very helpful and friendly thus far, checking in daily to make sure I have everything I need. He and Iryna are the two people I talk to the most. For the most part everyone does their own thing, though they are also relatively approachable. I will probably talk more about their personalities in a few weeks when I have a better idea of who they are. All in all, it's pretty comfortable in the lab, if also a bit individualistic. I see Dr. Fraser in passing most days. I will be sitting in my first lab meeting next Tuesday, which should prove to be interesting.
I think that should do it for today. In the coming days I'll talk more about some of the social events I've attended so far, getting to know the other grad students in my program(s) and stuff about Penn and Philly. I'm looking forward to what the summer will bring both in the lab and out. Hope you all are enjoying your summer!
Monday, June 15, 2009
Grad Diary 6/15/09
So, today was Orientation for those students opting to do a summer rotation. I chose to do one of my three lab rotations in the summer for a variety of reasons: a) I live relatively close to Philly, and so I can commute (for the summer at least), b) I would get my stipend sooner (and since I'll be commuting, save some major $), and c) I had nothing better to do for the summer, ha ha.
Anyway, as I mentioned, I'm planning on commuting to Philly for my lab work, at least for the summer. Once classes start, there is no way that commuting will be feasible, so I'll be moving into the city in August (almost done picking my apartment at the moment).
So I had today pretty well planned out; I had my map of campus, my train schedule, and a ton of nervous excitement to get me going.
I arrived in Philly at 30th St station a half hour before orientation started; plenty of time to catch the trolly (green line) to campus and find the meeting room. While I hadn't actually used the trollys (they go above and below ground in the West Philly area in addition to the traditional subway), I figured it would be fine, and I knew that the ride should only take about 5 minutes to go from 30th Street to 37th and Spruce, my destination.
I should note that, from 30th street to about 40th street the trolly runs underground like the subway system before returning to street level. My route would be entirely subterranean.
Boy was I in for an adventure.
As luck would have it (or rather, by Murphey's Law), the day I decide to take the trolly is the day that there are major technical difficulties as we make our way to the next station. The trolly in front of us was having mechanical difficulty, and to top it off, the driver was a newbie who didn't know what he was doing. As the minutes ticked away I tried to remain calm and refrain from constantly checking my cell phone for the time. I went over again and again in my mind where I needed to go so that, when I finally got off this god-forsaken contraption I could make my way quickly to my destination.
Eventually (30 minutes) we (literally) pushed our way to the next station...at 33rd street (yes, that's three blocks). At this point it was 9:00am, which was when orientation was scheduled to begin, and I quickly madem y way to the street level. No way I was going to continue on that nightmare. It took me a minute or two to figure out where I was exactly (I was smack dab in the middle of Drexel University) but I quickly figured it out and made my way to my orientation. I arrived there fifteen minutes late (that's right, it took half as long to walk the remaining six or seven blocks than it took to go three blocks on the trolley), and of course had to enter the door at the FRONT of the room, where everyone saw me rush in and find an empty chair way in the back.
In short, I was "that guy."
Great way to start my grad school experience.
A neat little twist to this story is that, since I came in late I wasn't able to grab my folder of orientation information, so I had to wait until there was a break to pick it up (at which point I apologized profusely). Before this, while the meeting was going on, another fellow arrived late and made his way to the back, sitting at the table next to mine. One of the administrative assistants walked over and asked for his name, then went ahead and retrieved his folder for him, while I remained folderless. The fact that this particular young man happened to be in a wheel chair probably had a lot to do with the differential treatment, which I thought was...interesting. Granted, I don't begrudge him for having different treatment because of his being differently abled - I am glad she gave him a hand, but it's not like I had any control over whether the trolly decided to malfunction that morning. This didn't bother me too much, but it did make me wonder about how we judge each other based on our outward conditions.
Anyway, the rest of orientation went fine, and I ran a few errands around campus for the rest of the afternoon (picked up my ID card, dropped some forms off, etc). I'll be starting my lab rotation tomorrow and I'm really excited, to say the least.
In the meantime, I have determined that I will be walking to work for the rest of the summer...
Take care!
Anyway, as I mentioned, I'm planning on commuting to Philly for my lab work, at least for the summer. Once classes start, there is no way that commuting will be feasible, so I'll be moving into the city in August (almost done picking my apartment at the moment).
So I had today pretty well planned out; I had my map of campus, my train schedule, and a ton of nervous excitement to get me going.
I arrived in Philly at 30th St station a half hour before orientation started; plenty of time to catch the trolly (green line) to campus and find the meeting room. While I hadn't actually used the trollys (they go above and below ground in the West Philly area in addition to the traditional subway), I figured it would be fine, and I knew that the ride should only take about 5 minutes to go from 30th Street to 37th and Spruce, my destination.
I should note that, from 30th street to about 40th street the trolly runs underground like the subway system before returning to street level. My route would be entirely subterranean.
Boy was I in for an adventure.
As luck would have it (or rather, by Murphey's Law), the day I decide to take the trolly is the day that there are major technical difficulties as we make our way to the next station. The trolly in front of us was having mechanical difficulty, and to top it off, the driver was a newbie who didn't know what he was doing. As the minutes ticked away I tried to remain calm and refrain from constantly checking my cell phone for the time. I went over again and again in my mind where I needed to go so that, when I finally got off this god-forsaken contraption I could make my way quickly to my destination.
Eventually (30 minutes) we (literally) pushed our way to the next station...at 33rd street (yes, that's three blocks). At this point it was 9:00am, which was when orientation was scheduled to begin, and I quickly madem y way to the street level. No way I was going to continue on that nightmare. It took me a minute or two to figure out where I was exactly (I was smack dab in the middle of Drexel University) but I quickly figured it out and made my way to my orientation. I arrived there fifteen minutes late (that's right, it took half as long to walk the remaining six or seven blocks than it took to go three blocks on the trolley), and of course had to enter the door at the FRONT of the room, where everyone saw me rush in and find an empty chair way in the back.
In short, I was "that guy."
Great way to start my grad school experience.
A neat little twist to this story is that, since I came in late I wasn't able to grab my folder of orientation information, so I had to wait until there was a break to pick it up (at which point I apologized profusely). Before this, while the meeting was going on, another fellow arrived late and made his way to the back, sitting at the table next to mine. One of the administrative assistants walked over and asked for his name, then went ahead and retrieved his folder for him, while I remained folderless. The fact that this particular young man happened to be in a wheel chair probably had a lot to do with the differential treatment, which I thought was...interesting. Granted, I don't begrudge him for having different treatment because of his being differently abled - I am glad she gave him a hand, but it's not like I had any control over whether the trolly decided to malfunction that morning. This didn't bother me too much, but it did make me wonder about how we judge each other based on our outward conditions.
Anyway, the rest of orientation went fine, and I ran a few errands around campus for the rest of the afternoon (picked up my ID card, dropped some forms off, etc). I'll be starting my lab rotation tomorrow and I'm really excited, to say the least.
In the meantime, I have determined that I will be walking to work for the rest of the summer...
Take care!
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