Mimbulus Mimbletonia

The DNA sequencing results all came back, so for the past two weeks, I've mostly been working on this program called GeneMarker that takes all of the DNA sequencing results and spits out the allele sizes at the EST/microsatellite marker loci as peaks on a graph. Here's a screenshot of a couple of GeneMarker graphs:

So the peaks portray how many base pairs long different alleles are. However, it's not as easy as it sounds because there are deceitful little peaks that try to trick you into believing that they're real - but really, they're just imposters... Anyway, I've been trying to tackle this program, but it's been putting up a good fight. But I'm determined to conquer it…somehow…

In the greenhouse, I'm still monitoring the germination and flowering of the M. tilingii and M. guttatus; however, many of the tilingii were looking slightly puny, so we ended up replanting some of them.

But thankfully, some of the paired guttatus individuals started flowering, and I began taking down morphological data. We decided on comparing and contrasting flowering time, basal width, overall height, number of vegetative stems, stem thickness between different nodes, internode lengths, leaf length/width, corolla length/width, stigma length, anther length, calyx length/width, and number of flowers after a certain time period.

We're also planning on measuring stomatal density and perhaps (hopefully, if all goes well) trichome density/morphology. Measuring these two traits is really interesting! So for stomatal density, you take casts of the underside of leaves using dental impression material and stick it on a microscope slide. When that dries, you spread some clear nail polish on the casts and make leaf impressions from the nail polish peels. You put those peels on another microscope slide and examine it under a microscope, and you can see individual plant cells, including the guard cells that control stomatal opening/closing! Super cool! Also, I've been wanting to examine more closely the tiny little hair-like structures that are found on both the tilingii and guttatus. Here's an amazing picture of a tilingii individual in which the trichomes are really easy to see (taken by Carrie):

Amazingly, some of the tilingii populations (like this one) have a sticky substance on their trichomes. It's hypothesized that this glandular exudate is a mechanism for resistance to insect herbivory, insulation, etc. Anyway, we looked at the trichomes on both the tilingii and guttatus and discovered that for the individuals that we examined, the trichome morphology was different between the two species

Oh, and I almost forgot! I was reading Harry Potter and the Order of the Phoenix, and there's this part that describes a plant that Neville has called "Mimbulus mimbletonia". Doesn't "mimbulus" sound eerily similar to "mimulus"? Anyway, I looked it up on Wikipedia:

"Mimbulus mimbletonia is described as "appear[ing] to be a small grey cactus in a pot, except that it was covered with what looked like boils rather than spines" (OotP 186). The genus name mimbulus may be related to the real genus mimulus, especially since those plants are used as folk remedies for "shyness, anxiety, and forgetfulness" and those are traits of Neville Longbottom."

Mimulus are definitely not small grey cacti, but I thought that was a cool coincidence. Now whenever I go to the greenhouse, I feel like I'm in Professor Sprout's class! =]

Plant Nerd

At the beginning of the week, Carrie showed me how to set up a 96-well tray for PCR with single EST markers. I found that it's really hectic when I'm making multiple cocktails, each with a different primer, and then realize that I'm running low on Taq, primers, dNTPs, or buffer...all at once. Anyway, I actually finished the PCRs with all of the primers that I'll be using in my project, but I still have the microsatellites to do. With the PCR results, I ran gels and found that most of them all had bands, which made me really happy! Once, when I was microwaving the agarose + buffer solution, I got caught up in talking to someone and forgot about the beaker! So the solution spilled over... but thankfully, it wasn't anything serious. I remembered that safety video and told one of the lab people what happened (even though I was completely mortified), but she was really nice and told me how to clean it up =]. After checking for the germination of the remaining seeds, David and I collected seeds from plants that had fruited and selfed those that hadn't been crossed yet. During that time, I had a chance to ask David about his "life path", and I found out that he had a fascinating story to tell. Then up in the lab, I went to Google Maps and found the locations of all of the tilingii and guttatus populations that I'm studying, and I made my own map for my poster. Oh, speaking of posters - today, we dug out old posters that various people from the lab had made, and David showed me how there is a variety of ways to set them up. Along with that, I heard/read about the fascinating research of past and present lab members. It's been a busy but fun-filled week...

I've been thinking - I've always considered myself an animal-person (my house = perpetually overcrowded zoo / animal hospital), and I am...but this summer, I realized how incredibly amazing/intruiging/unbelievably complex plants are! Since I was young, my parents bought flat after flat of plants from Lowe's and also rescued unwanted ones from various people moving back to Japan, and I had always wondered what it was about plants that fascinated my parents. I viewed plants as boring because no matter how hard I worked, they never seemed to interact with me - they don't play fetch, hop onto my lap, beg for carrots, or curl up in my hair. But I realized this summer that plants aren't just those green, crunchy, photosynthesizing "things" that we eat in salads - they are incredibly diverse and have so many different mechanisms for survival that have evolved throughout their history. I'm kind of disappointed about not realizing this earlier...It's kind of like when you're in the same class with a person for a whole year, but you never really get to know her until the last day when you realize how charismatic she truly is...and then you want to hit yourself for not taking the opportunity to get to know her better when you had had the chance to for the whole year. But I'm happy because I have the rest of my hopefully long life to get to know plants better.
I can't believe it's already been two weeks! I've now begun stopping to examine random plants while I'm running and looking at the distance between the anther and stamen (I'm really interested in that trait!), and I'm now performing experiments with the various begonias, Saint Paulias, and Torenias around my house to see if I can cross-polinate and/or self them. I feel like such a plant nerd...and I love it!

First week with the monkey flowers

My name is Mai Nakamura, and I'm a rising senior at East Chapel Hill High School. My PI for the program is Dr. John Willis, and I'll mostly be working with Carrie Wu and David Lowry. My project will be to figure out whether the Mimulus tilingii is more similar, in terms of its DNA and its morphology, to the geographically neighboring Mimulus guttatus or more similar to other Mimulus tilingii that are farther away. I think this method of working on two projects at the same time is wonderful for me because I not only have the opportunity to work with the molecular component of Mimulus but I also don't lose track of the "big picture" by actually doing physical gardening-type experimental work and getting soil in my fingernails which allows me to keep in touch with the underlying goals of the molecular work in the lab. I don't know about the rest of you but for me in biology lab last year, I would often know to stir this enzyme into this test tube or to pour a dye into that cocktail, but I didn't have a strong grasp of what I was supposed to be trying to accomplish. There would always be a big, gaping question - "Why am I doing this, and what is this for?". So I really like this method of two ongoing projects because it answers this question and doesn't leave me in a state of uncertainty =].

Monday: I had already met most of the people in the Willis lab last week, so Carrie and David immediately started me working on relocating various pre-planted Mimulus seeds that had been going through "winter" to the greenhouse where they will hopefully start germinating in the next week or so. With a clipboard, I wrote down the population names and the individual's number on a sheet of paper, and I later put that information on an Excel document so that the recording of the germination/flowering dates will be easier later on. Then, David and I went back down to the greenhouse and started "selfing" or self-fertilizing some Mimulus guttatus. (I found out that it's a lot harder than it looks/sounds...) Then, we went up to the lab conference room, and David and Carrie began to explain to me the concept of QTL mapping. I'm really lucky to have two mentors that are both really good at explaining hard concepts!

Tuesday: After this morning's gel electrophoresis, I went to the lab and did gel electrophoresis again with Carrie! It was great because I actually felt a lot more comfortable the second time with having this process still fresh in my mind. Hopefully, it'll be drilled into me by the time I have to do it for actual experimental data. So for the gel electrophoresis, we used some PCR results from another lab member and a hyperIV ladder...And it worked out well! We used the machine with the ultraviolet light (I don't really know its name or how it works) and printed out a picture of the gel.

Wednesday: Amazingly, some of the seeds began to germinate after only two days! However, seeds from some populations, probably those that have a long growing season that can afford to germinate later, remained dormant. The other major event of the day was "PCRing" with Carrie. It was a lot harder than I thought it would be, with all the different programs depending on the microsatellites used or whether ESTs markers are used. At the end, thankfully, all the tubes seemed as though they had about the same volume of liquid in it, so hopefully the gel that we will be making tomorrow will show good results.

Thursday: Today, Carrie and I used gel electrophoresis to test to see whether we had good PCR results from yesterday. Carrie pretty much let me make the gel by myself, always standing right next to me and pointing a few things out once in a while. While we let the gel dry, we went into the conference room again, and Carrie went over in detail what she would like for me to accomplish this summer. Since Mimulus tilingii are high-altitude plants that live all the way up in the alpines, researchers don't yet know whether they evolved independently from convergent evolution or whether there had been a widespread area characterized by cold weather way-back-when that eventually diminished to create these now isolated Mimulus tilingii populations that can only survive at high altitudes. So, what I'm going to be doing this summer is comparing, phenotypically and genotypically, the Mimulus tilingii populations among each other and between Mimulus guttatus . If we find that the Mimulus tilingii are more similar to each other, this probably means that the wide-spread colder region did exist and that there was more recent gene flow among the tilingii . On the other hand, if we find that the Mimulus tilingii populations are more similar to the various Mimulus guttatus populations, then that suggests that there has been more recent gene flow between the two separate species populations. After that, we went back to the lab for the running of the gel after which we discovered that the PCR had gone well! Once again, using that UV light machine, we printed out a picture of the gel.

Friday: Today, we did another PCR but with two different markers - EST641 and EST332. While the PCR was going, I made various dilutions of all the markers that I will be using along with some dNTP. After, I made another gel with the PCR results, and I saw that I hadn't done as well as the previous time...but it wasn't bad. That pretty much took up most of the afternoon.

I hope everyone else is enjoying themselves as much as I am! I am absolutely exhausted...