Monday was a crazy day.
So I had to sign up for lab safety training because I’m working in a wet
bench lab. So I registered for the
course and signups were at 9 and the actual training began at 9:15. Unfortunately, my mom and I couldn’t find any
parking and minutes were ticking by. Finally
we decided to just park in one of the decks, but it was a few blocks from the
address they gave us. Anyway we parked
and walked down the street. Once we got
to the building we couldn’t get in the one side so we had to walk to the other
side and finally got in. The man at the
desk told us that this was not the right location and it was down the street
where we had to go down an elevator to the street below the one we were
currently on. We found where the
elevator was which was in a glass room on the side of the road. There was another desk that directed us to go
outside and the entrance was on the left.
We reached the door at about 9:30, after all our troubles it was a
relief. We read the sign and rang the
bell. Turns out the doors closed exactly
at 9:15 and they weren’t letting anyone in.
It was a very frustrating morning.
I got to the lab at 10 and started working. It was an RNA day. Since RNA is denatured by RNases, which are
on our skin and on our clothes because it could be harmful if it somehow entered
a cell, Julie and I had to wear lab coats along with our gloves. There is also a special bench for working with
RNA and everything has to be sprayed with RNase ZAP and you have to be cautious
or the RNA will be denatured and therefore useless. We had to add chloroform to the TRIzol, which
separates the RNA, DNA, and protein, and then centrifuge them. A hard part was extracting only the layer
with the RNA, once that was done, isopropanol was added to help the RNA
precipitate from the solution when centrifuged.
When that was done the pellet was not visible but we dumped out the supernatant
and then added ethanol. After that was
centrifuged, you could see a pellet in some of them but not all and this time
we had to get all of the ethanol out.
That was tricky but we let them dry to have as much ethanol evaporate as
possible. We added water and then placed the tubes in
the water bath to resuspend the RNA. To
determine the RNA concentrations in each of the samples, I used the nano drop
and this time I used the other part of it where it’s a small metal piece
protruding from the surface and a small amount of the sample is placed on it
and the machine reads the concentration.
Unfortunately, the numbers were a little low for RNA but we decided to
run PCR anyway. Before I knew it my mom
was here.
Tuesday began with a nice ride with Dr. Peretz. Since she is coming to visit the labs, she
offered to drive anyone that needed a ride and since my parents have a funeral
to attend, it worked out nicely. We
talked the whole ride about various things and it was very enjoyable. I got into the lab and since Julie was out, I
thawed the primary and put it on one of the blots we used last week. While that sat in the cold room, I listened
to a podcast on CRISPR since that’s what we’re working with. Then we had our group lunch. Julie and I started walking over to the vet
building where Matt and Amber are working.
As we walked, we passed Dr. Peretz who was driving us over to the Shake
Shack. Once everyone gathered in the car
we drove over and then ordered. The
whole lunch was a lot of fun, seeing everyone and talking about our experiences
was very interesting. We got back and
Dr. Peretz came and visited the lab, she asked questions about what I was doing
and the overall project. Then I finally
got a picture in a lab coat! Her visit
was really nice and being a bio teacher and a geneticist herself it was in her
territory. Then, when she headed out
Julie and I planned out the plate for the PCR.
It got difficult to plan out where everything was going so that we could
fit what we needed onto the plate.
Finally we got that all figured out and started to make the primers that
needed to be with the RNA in the plates.
Once all that was mixed, Julie went to the tissue culture room to plate the
cells we had just received. It was
perfect timing because as soon as the plating was done, Dr. Peretz was pulling
in front of the building. Unfortunately,
it was pouring when I went outside but I ran through the rain and got in the
van and headed home.
Wednesday began with filling the wells with the cDNA and the
primer to get it ready for the PCR machine.
Cagla helped us set up the machine to make sure it was going to run
correctly. That was left for two hours
and while that was replicating I was with the microscope taking images of the cells
I had treated last week. Unfortunately, the
PSD95 didn’t work great on those cells but the MAP2 and the GFP did so I took
pictures with those fluorescing. In
total I took 130 pictures and tried my best to look for the neurons that had
the clearest neurites and where the neurons weren’t on top of each other. I grabbed some lunch and waited until Julie
got back from her lunch. We started working
with the western I had put the primary on yesterday. This time we did four 10 minute washes to cut
down the time so that we could go to the Happy Hour celebration for the
lab. Kelly got a huge grant that she had
been trying for a while and so when she received it, it was very exciting so we
are going out and this is also a farewell to Julie because tomorrow is her last
day with the lab. When she leaves I will
be continuing the project on my own but I’ll have Cagla and Shelly as well as
Eddie and Anna if I need anything or I’m confused. I finished with the blot and the film I
developed was actually better than a lot of the ones we previously got. We headed over to Mad Mex for the Happy Hour
celebration. Unfortunately, I wasn’t
able to spend a lot of time with the group but it was still enjoyable being out
of the lab with the group.
Thursday was a busy day.
Since it was Julie’s last day, she wanted to make sure I knew where
everything is and what I will need when she’s gone. I also brought in donuts for Julie’s last day
which ended up being a very random assortment with some donuts I’ve never seen
before. Anyway, we began our day by
looking at the neurons we plated on Tuesday and seeing how they’re coming along
and heated up the media. Then the media was placed in wells with the treatment
we have been looking at in other experiments.
We also began more washes on the blot I used yesterday in hopes of
getting a better final result. We added
the treatments to the cells and had them sit on this one machine that will pull
the magnetic beads through the cell. And
we started with the treatments for another plate as well. We put those on the cells and placed the
cells on the magnetic plate again. I put
the secondary on the blot and took the cells off the machine when they were
ready and set them in the incubator. We
had time so I relaxed and wrote some of this as well as read my summer reading
book, The Lords of Discipline. Julie and Brigid, another grad student in
the lab, along with Shelly took a potential member of the lab out to
lunch. I stayed and continued working on
the blot. I did the washes and
everything went very smoothly. I headed
down to the dark room and met up with Jenna, an undergrad in the lab, who was
also down there. Julie met me down there
when she got back from lunch and our results ended up being better than we were
expecting so that was exciting. Once we
finished scanning those blots in Julie was showing me everything around the lab
that I will need for my remaining time here.
Luckily I have done almost everything that I will do and therefore have
at the least an idea of what to do, but like I said I will have everyone else
in the lab if I need to ask questions. The
end of the day was quiet since we finished with much of what we had to do and
Julie was clearing her stuff out while I was writing this and reading. My mom finally arrived after hitting a lot of
traffic. I gave Julie a hug because she
is an amazing person and has done so much for me in these short four
weeks. I am so happy I got a chance to
meet her and work with her.
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