Institute of Regenerative Cures

My class got to go on a field trip last week.

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All the joys of barely being able to see the tour guide when you’re at the back of the group.

I know! Field trips in a Master’s of Science program? How ridiculous!

It was awesome. We went to the Institute of Regenerative Cures in Sacramento.

I arrived early and waited out front with some classmates. Our tour guide arrived and we waited out front a little longer til everyone showed up.

While waiting the tour guide, who had designed the building we were about to go into, told us about his hobby, early television history!

After the primer on early television we entered the building and got a tour of one of the best facilities for practicing biology in existence right now.

The building itself was actually built a long time ago for the California state fair. It was the “women’s building.”

The brick exterior and columnaic entrance have stayed the same since the building was constructed to maintain the historical site. The interior has been heavily modified.

The building had no roof back in the day and was just an enclosure for a bunch of different events that you usually see at state fairs.

The building was sold to the University of California system. They slapped a roof on it, and used it to store records.

Our tour guide said that he was called in to turn it into a biology facility later on. Half the building is used for bio research while the other half is rented out to other companies.

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The entrance hallway with pictures of the cooler discoveries at the Institute.

The researchers in the Institute are working on a number of things. They researched a treatment for the “bubble boy disease” there. They’re working on using umbilical cords to create bone marrow for transplants, using Tal proteins to treat Huntington’s, creating HIV resistant cells, and helping people who can’t swallow to swallow are just a few of the things they work on there.

Where all the research is done!
Where all the research is done!

The tour guide also showed us the section that he was most proud of as he had designed it. A set of rooms for making the actual drugs and proteins to export to hospitals. Making the drugs requires extremely sterile technique to prevent giving someone who is already sick something that will make them worse. The rooms are designed to be extremely sterile.

To enter the rooms you pass through an airlock where you are required to cover every inch of your body in a disposable gown.

The airlock goes to a hallway with access to three separate clean rooms.

There is “negative pressure” in the rooms. That means that air is constantly entering the room from the top and going out the bottom. This is so that if any cells that are worked with in the rooms get into the air, they will be redirected to teh ground and sucked out through a grate in the wall instead of ending up in someone’s medicine.

The air is cleaned excessively to about 3000 times more clean than average air before entering the facility.

There is a lot of electrical equipment in the rooms that will require replacing eventually. To prevent electricians from having to gown up just to replace a lightbulb, all the eletricals are accessible from panels on the second story of the building.

It was pretty cool for a scientist like me to see the best possible place to do research in. The tour guide mentioned that he does tours of the interior of the super clean rooms for smaller groups. I might take him up on that at a later time!

-GoCorral

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My Master’s Project

All labwork is overseen by the disembodied head of Muppet lab assistant, Beaker.
All labwork is overseen by the disembodied head of Muppet lab assistant, Beaker.

I have begun my Master’s project in earnest and the goal is slightly different than what I’d been doing before.

First, I’ll repeat myself. I’m a biologist and I work with introns in C. elegans. C. elegans is a type of nematode worm that naturally lives in soil or on rotting vegetables. It is also one of the most widely used model organisms for biological research.

Worms! Ew! Gross!
Worms! Ew! Gross!

Introns are unused sections of genes. You’re probably aware that DNA is in our cells and contains the instructions for how an organism functions. The human genome contains around 25,000 genes and those genes are split into two parts, introns and exons.

Exons are the part of that gene that are actually used to produce things in your cells, while introns are spliced out and removed. So why are introns on there at all if they’re removed?

Well it turns out that some introns increase expression of the genes they’re in. My project looks at how placement of those enhancing introns affects expression.

Experiments in plants have shown that an enhancing intron works best when it is placed near the start of a gene. Experiments in C. elegans have suggested that, but no experiment has outright proved it. My project will hopefully do that.

I’m measuring the expression of genes according to how introns affect them, so I get to pick which gene to use. When picking a gene like this scientists often pick what are called reporter genes. The expression of these types of genes is easy to measure, often because they have produce light or fluorescence of some kind. The light tells you whether the gene is on, but also at what level it is turned on based on how bright the cell is.

Previously I was using a reporter gene called GUS. GUS is an enzyme that digests a specially prepared sugar, releasing a blue chemical that was attached to that sugar. The blue chemical is then visible to the naked eye.

There were a number of problems with that experiment though. First, adding the sugar chemical to the worms was a pain, taking about three days to set up and look at. Plus, the blue color was difficult to measure precisely because most of the machines in the lab are set up to measure red or green colors, not blue. Finally, GUS is traditionally a reporter gene for plants, not C. elegans. This could’ve been introducing other problems that we couldn’t easily identify. Thus the use of the GUS reporter gene has been scrapped in favor of another reporter gene.

I’ll be using Green Fluorescent Protein (GFP) as my reporter gene now. GFP is widely used in C. elegans and many other organisms. The protein created by the GFP gene glows green when you shine a red light on it. Very easy to see and measure. None of that three day procedure for GUS. I just pop the worms under the light and take a look.

Why weren’t we using this procedure before if it’s so easy? Two reasons!

Reason number one: C. elegans won’t express GFP without introns in the gene. So does that mean we proceed and hope one intron is enough or do we add the standard amount of introns to get expression? I’ve decided to see what the GFP looks like with the standard introns scientists put in it for C. elegans and without them. I’ll also be testing with an added intron. The whole thing is a little complicated so here’s a diagram to explain.

Here are the constructs I've been creating. The wide parts are exons and the thing parts are introns. The green bands are the GFP which will glow green in the worms. The white bands are a scaffold which allow the worms to express GFP
Here are the constructs I’ve been creating. The wide parts are exons and the thing parts are introns. The green bands are the GFP which will glow green in the worms. The white bands are a scaffold which allow the worms to express GFP.

There are eight different constructs I’m making. They are a combination of three different features that are present or not. Are there introns in the first GFP? Yes or no? The second GFP? And is the Unc54 intron there? This allows us to control for the positional effect the standard introns in C. elegans GFP.

Reason number two: Those eight constructs above? Those aren’t made yet! All the GUS constructs were made when I started the project. I’ve been working on making the new constructs for a few months. It could take a few more months to finish.

So my project is to make those constructs, put them into worms, and then see what the worms look like. As I perform these steps I’ll make more posts about what work I’m doing in lab and why its so cool.

-Mister Ed

TAing at Sac State

One of the lab benches of the room I teach in.
One of the lab benches of the room I teach in.

For the past few months I have been assisting in teaching a introductory biology lab at Sac State.

I TA Bio 15L which is a general education course for non-science majors. The course uses interactive labs to go over all the basics of biology, like ecology, speciation, DNA, genes, and that good stuff.

The class has been a lot of fun for me for a lot of different reasons.

I like helping out the students. Its nice to see some of them so interested in biology even if it is nowhere near what their major is. One of them is even considering switching her major.

It’s nice to go over all the material again. I learned it all years ago and everything is easy for me now. Obviously I should know the material in a class that I teach, but its still fun to know that I could get any of the questions in the class right if the teacher called on me, even when I am the teacher.

The experience of being on the other side of a class is also interesting. I have to deal with making quizzes, grading, student absences, and preventing cheating.

Student absences is probably the hardest part. This is a college level class, so they’re free to not show up if they don’t want to. Its just inevitable that the ones who don’t show up do poorly on the quizzes that cover the material they missed or they miss the quizzes entirely. And this is college so there are no makeup quizzes.

There’s nothing I can really do about absences, but I’d like to be able to tech the students that do come to class so that they can all understand the material and use it in their own lives later on.

Learning biology is important for a number of reasons. How can you be an informed voter on GMO issues if you don’t properly understand what GMOs are? How can you vote on global warming initiatives without knowing more about that? And wouldn’t you like to know how genetics work when you start planning a family to see what genetic risks your potential child could have?

I try to teach the students that sort of stuff. I feel like I’m just learning how the labs work this semester. I know I’ll do way better next semester when I can focus more on directing what we are trying to learn with the labs and giving the students more specific strategies for learning.

Also, I can hopefully be more enthusiastic when I give lectures. The mid-semester student evaluations indicated that the only place I really needed to improve was in how enthusiastic my voice sounded when I was presenting the material.

-Mister Ed

Which Lab for Grad School?

This is the microscope I use to inject DNA into nematode worms.
This is the microscope I use to inject DNA into nematode worms.

I’ve been doing some thinking lately about which lab I should work in for grad school.

As it turns out I get to choose among a few different options.

The folks at Sacramento State are okay with me doing my research at either of my labs in Davis.

I’ve been with the rice lab for almost three years now and feel I’ve gotten what I wanted to out of it.

I’ve already written some goodbye/thank you letters, but have yet to hand them out. I’m just ready to leave the rice lab.

Yesterday I looked up some information on what exactly I’d be doing if I joined the new professor’s lab at Sac State.

The professors old students finished their theses which are then stored in the school library.

Recently the library has started putting digital copies of the theses online. I read a few of the more recent ones that were uploaded.

While the research is interesting, there was nothing that I wanted to do more than the intron research I do currently.

Part of it was the occupational hazard of working with food pathogens. Most food pathogens are collected from raw food samples or from poop.

The idea of having to collect poop samples and work with them… Let’s say its not on my bucket list and leave it at that.

Continuing my intron research would be awesome though. The project has room for expansion and it fits better with what I want to do on a grander scale.

I want to create tools for people to use in other laboratories. Enhancing introns could be used in any laboratory to fine-tune the expression of a gene to the exact level required for an experiment.

I want to create tools like that when I get an official job as a researcher, so it would be best if I did my Master’s Thesis on the same topic.

So it looks like I will be attending Sac State next year but performing my research at UC Davis on introns!

-Mister Ed

Disposing of GMOs

The rice we grow in one of my lab's greenhouses.
The rice we grow in one of my lab’s greenhouses.

Yesterday I was working out at the greenhouse for my rice genetics lab.

I was getting rid of some old rice plants that we’d collected the seed from and no longer needed.

If a plant got to this point in a garden you’d normally throw it in the compost so it would be useful next year.

That’s not allowed for the rice we work with in my lab because it is an untested transgenic line.

Some members of the public dislike altering the genetics of food crops to create genetically modified organisms (GMOs). There are a couple of logical reasons for this and a couple of illogical ones.

Logical reasons include: religious objection, lack of crop diversification, cross-species allergens, and the strengthening of agribusiness monopolies that often accompanies GMO crop use.

Illogical reasons often have something to do with safety or not knowing what is in a product when you purchase it at the grocery store.

I could go on about this a lot. GMOs are a complex topic with a lot of ground to cover, but that wasn’t why I was writing this post today.

Because of the fear of GMOs, they need to go through extensive testing before they are declared legally safe. This testing can take up to ten years.

We don’t do that for every strain of modified rice in our lab, so certain precautions need to be taken.

Yesterday I cut off all the excess seeds on the old rice plants. The seeds go into a plastic bag.

The seed bag and the leftover portion you can see above both go into an orange dumpster at the center of the greenhouse complex.

All the stuff in the orange dumpster then goes into a special oven that ensures the modified crops won’t somehow get into the wild and start growing there.

After the special oven, called an autoclave, has destroyed the genetic material in the rice it can go into a normal dumpster or be used for compost.

Just another little glimpse at my job!

-Mister Ed

Coke Floats and a Full Day

Two Coke floats my wife and I made.
Two Coke floats my wife and I made.

I wanted two root beer floats tonight, but we didn’t have any root beer in the house.

I made Coke floats instead.

We only had enough vanilla ice cream for one float, so I used chocolate in the other one.

They were not really root beer floats, but they were really nice.

And apparently my wife never had a root beer float, so this was her first experience.

The floats were an end to a long day which involved: getting fewer privileges at work, trouble shooting a problem at work, talking about relationships, having a birthday lunch with a friend, organizing vacations, doing laundry, cleaning cat poo, and at the end I got floats!

Previously I had thought it was alright to watch Youtube videos at work while I sorted seeds. Boss said no today. I’m disappointed, but its not unreasonable of him to ask this.

Isolating DNA from rice leaves hasn’t been going well. We’ve been getting errors and we finally narrowed down the error to a salt we use.

The stock salt we use to make the salt solution looked dirty though. I cleaned it. We’ll see if it’s good next week.

I talked with my coworker while sorting seeds about his future plans with his girlfriend. He wants to move in with her, but she thinks marriage is the next step.

They’ll probably just live close to each other next year. Right now they live around an hour apart.

One of my old friends from my organic chemistry class sophomore year had a birthday today!

I took him out to lunch at Taco Bell (he chose it). We talked about life and stuff. It was a lot of fun!

When I got home I organized my Memorial Day vacation with my father-in-law.

Another old friend, from high school this time, wants to hang out on the actual Monday of Memorial Day weekend. Organized stuff to make it work.

Laundry and cleaning the litter box are pretty typical chore things.

And chocolate ice cream in vanilla Coke to end the day!

-Mister Ed

Injecting Worms

This is what my computer captures under my microscope when I inject a worm.
This is what my computer captures under my microscope when I inject a worm.

I gave an extra post about one of my jobs. It seems fair to cover the other job as well at some point!

I study introns in C. elegans worms, but how do I get the specific introns in the worms?

I need introns in specific placements in specific genes in order to study them with scientific accuracy.

The gene we are studying is simple. If the worms are put in a solution called X-gluc, they turn blue.

Based on where our enhancing intron is in the worm we expect it to turn more blue if the intron is closer to the start of the gene or less blue if it is near the end.

So I have these genes that I’m putting into the worm. They get in by injecting them like you see in the picture.

The needle of DNA is aimed at the gonad of the worm.

C. elegans worms are hermaphrodites. They contain sperm and eggs and they self-fertilize.

The worms are “male” at first, producing a bunch of sperm.

Later on they produce eggs and then they fertilize their own eggs with the sperm stored in their body.

Since they contain both genitalia the whole area is referred to as the gonad.

I aim my injection at the gonad, hoping that the DNA I’m injecting will get into the fertilized eggs.

Then the injected worm is put on a plate with lots of food and I hope that its babies will have the injected DNA.

But I don’t test for “blueness” immediately.

When a worm is first injected, the DNA is inside its cells, but not necessarily integrated into the cell’s chromosomes. I need the DNA to be a part of the chromosomes.

There are only two genes in the mix of injected DNA that will integrate. One gene is the blue gene, called GUS. The other gene is called unc119.

Unc119 is to “recover” the worms.

The worms I inject lack unc119, which is a normal gene for worms.

In a natural wild-type worm unc119 aids the development of the worm’s neural network. Without it, the worm has poor neural connections and has a lot of trouble even moving around and eating.

So the first way I test a successful injection is by looking to see if the babies of the injected worm are moving around normally or flopping around.

The normally moving ones were successful and now have unc119. They are “recovered” back to their natural wild-type state.

The floppy crippled ones did not have a successful injection. Either I missed the gonad, I didn’t inject enough DNA, or the eggs that got my injection didn’t fully germinate.

There are other markers I use to see if an injection was successful, but I’ll get to those later!

-Mister Ed