Technology in Medicine (Sci-fi or Reality?)

Today, we learned to adjust and use micropipettors, which are used to measure very small amounts of liquid very precisely. We then used these skills to transfer genes to E. coli bacteria. If our procedure worked properly, we should have fluorescent and antibiotic-resistant bacteria by tomorrow.

Our materials for the gene transfer

We practiced using micropipettors to place small amounts of liquid on the sides of microfuge tubes to prevent them from mixing before they should.

We used nichrome wires to spread bacteria on agar inside the Petri dishes.

Sterilizing the nichrome wire with a Bunsen burner.

Today, our class started off with a Skype call to Dr. Kwong, an aspiring oncologist and current trauma surgeon. Dr. Kwong shared her plentiful knowledge of surgical oncology, how to become a doctor, and trauma surgery. One particularly interesting thing she discussed were the different cases that came into the trauma rooms from day-to-day. Because there is no set time or pattern for traumas to occur, she could be called in and only have one patient , yet she could be called in the next day and have seven to ten patients. When asked what her most interesting trauma was, Dr. Kwong said that once, she had two patients that were shot 6 times. Although one died, the other came out with just a broken toe. The wide array of patients and cases was very interesting to hear about, and exciting to learn about.

After our break, the class practiced sutures on bananas. We watched a video first to learn how to use the tools we had and how to hold the skin together. We opened our bananas, removed the actual fruit part, and started trying our individual stitches. The whole class got better with every stitch we tried, and by the end, we had closed all the bananas all up. After lunch, we retrieved our petri dishes from the incubator and inspected the colonies of bacteria that had grown. Our group was very successful and had plentiful amounts of colonies that formed and that glowed in the dark.

Today, we traveled to the Griffith Lab at UNC Chapel Hill, where we watched a presentation about the research being done at the Griffith Lab regarding the use of electron microscopes with DNA. We learned about different methods of preparing the samples for viewing under the electron microscopes including thin sectioning. This method involves cutting the specimen into thin slices in order to be viewed under the electron microscope. We were able to look closely at the diamond knife which is used to slice the sample. We also had the opportunity to observe and use a transmission electron microscope and watched how they loaded the sample carefully into the microscope and use various knobs and levers to focus on a specific part of the specimen while the electrons were shining on it. We also saw how you are able to view the specimen either by looking through an eyepiece in which you must switch a lever to see through or on a screen that was connected to a camera attached inside the transmission electron microscope in which you must switch another lever to allow the camera to see. The trip to the Griffith Lab was a great experience that we are very thankful for and in which we were able to see how researchers use and apply equipment that we have learned about in the field.

he most interesting thing that happened during the dissection was examining the heart and lungs of the rabbit.  We had some difficulty cutting open the chest cavity of the rabbit without damaging the heart and lungs of the rabbit.  We started cutting open the rabbit too low at the liver instead of the just below the ribs.  This caused us to have to cut through the ribs to expose the heart and lungs.  I thought that it was interesting how small they were in the large rabbit.  It was also interesting that the heat and lungs were small, because rabbits hop very quickly and are quite fast.

 

The most difficult part of the dissection was determining the best way to make incisions and cuts on the rabbit.  When tanning the rabbit we did not want to cut into the muscle of the rabbit which was why it was so difficult.  We also had some difficulty cutting open into the chest cavity of the rabbit without damaging the heart and lungs.  Identifying the body parts and organs was also difficult, especially for people who have no prior experience.  Beyond that, clean-up was difficult due to all the hair getting everywhere.

 

The most surprising part of the dissection was how easy it is to remove the skin of the rabbit. With the fetal pigs, my group had some difficulty getting into the pig. We had no such difficulty getting into the rabbit.  The connective tissue was very easy to remove and only when there was some fat between the skin and the muscle was it somewhat difficult.

 

We are looking forward to dissecting out the rabbits heart and closer examining it.  This will be very interesting because we were unable to do this when dissecting the fetal pigs.  We have previously learned how the heart works and how blood flows through the heart, so it will be very interesting to see what the heart look like in person.

 

Here are some photos of our rabbit dissection:

 

We also talked with Dr. Dienst about his experiences as a medical doctor. It was interesting to learn about how much medicine has advanced in the past few years.  According to Dr. Dienst, ~70% of the current medical procedures were not being performed when he was first a doctor.  In order to keep up with new medicine, he said that he reads six or seven articles per day.

One of today’s objectives was to suture our rabbits, but we couldn’t just start on the rabbit without practice. Hence we practiced on banana peels before lunch. First we removed the banana out of its peel. With the peel an incisions were inflicted upon with a scalpel to close up. To make sure that we didn’t cut ourselves with the needle we put on gloves and used clamps. We practiced individual knots and running stich. Running stich is quicker to do, but it causes more trauma and visible scars. It is different from individual knots, because string continues instead of knotting individually and cutting it. Individual knots are more common since there would be minimal amount of trauma and scars. 

This morning, we went to Dr. Griffith’s lab.  Interestingly, Dr. Griffith is my piano teacher’s husband, so I already knew him.  We first watched a presentation about thin sectioning, DNA, and viruses, and how they relate to the electron microscope.

It is amazing how powerful and expensive these tools are, and it’s simply unbelievable that we can now see what we’ve never seen before.

We also got to see the diamond knife. Dr. Christina showed us the way it works, then put it under a microscope with a specimen, and let us look into it (it had two eye pieces!!!).  The diamond knife cuts specimens for thin sectioning.

 

We started off the day by facetiming Dr. Kwong about cancer, trauma, and the process of training to be a specialist. It was startling how different each cancer case is.  One thing I learned is how cancer is staged – 1-4, depending on how far it’s spread.  Next, we watched a video on suturing and practiced suturing on bananas using braided silk thread pre-attached to needles.  We practiced two types of sutures: interrupted and running.

After lunch, we looked at the results from the DNA experiment.

#	Contents	– : no amp	+ amp	++ amp + arabinose
1	control – pGLO	a lot	confluence	confluence and they glow!
2	control: UC19	n/a	confluence	n/a
3	Ligation: GFP	n/a	none	none

Table 1. The results of the DNA experiment

 

The reason we didn’t have anything from tube three is that something went wrong during the ligation reaction – the error was not in the transformation, because the controls still worked.  Even though tube 3 didn’t work, tube 1 in the ++ plate was very successful and glowed strongly.

Welcome to the blog of team Scrubs: Alexandra and Amy.

Today we practiced micro-pipetting. There are 4 types of micro-pipettes that we used: P 0.5-10, P 2-20, P 20-200, and P 100-1000.

P 100-1000, P 2-20, P 20-200:

P 0.5-10:

We first practiced on colored water to make sure that we were ready for the real experiment we were about to do.

This is what our sheet looked like:

We used disposable tips to make sure there was no contamination.

YAY! COLORS:)

We did a mini lab where we put the color on the side of the tube to practice our skills.

 

Then we moved on to the real experiment!!

 

Enjoy the movie below!!!!!!!!!!!!!!!!!!!!!!!!

Here is a closer picture of the diagram of the cell we were genetically modifying:

We’ll catch you tomorrow! See you then!

~SCRUB OUT~

Thursday morning, we began practicing our micro-pipette skills by pipetting colored water onto a square of plastic. We had to first practice measuring how many milliliters we were dropping by moving the wheel on micro-pipette. To finish off the morning, we made stain glassed pieces by micro pipetting different colors into a 96 hole plate. After we had lots of practice pipetting, we conducted an experiment that resulted in E.coli colonies that were antibiotic resistant as well as glow in the dark.

This is a video of Jaida ejecting some colored water from a Micro-pipette. Unfortunately, her hand was a little shaky.

DT- shaky pipette-25tz9m2

Today, we worked to insert the gene that makes jellyfish glow into the E. Coli bacteria. As this process involved many elements of using micropipettes, we first began with practicing our micro pipetting skills. We did this by performing various exercises, one of which involving creating micropipette art! In one of the exercises, we also used a micro centrifuge to centrifuge the colored liquids which we had pipetted on the sides of the tube together.

The first step in the gene injection process was to perform a ligation reaction with three tubes–two being controls and one meant for the bacteria. The next step was transformation in which we were required to create sterile environments at each of our stations, in which we were actually dealing with the bacteria. This process consisted of many intense and time dependent steps including thawing the bacteria on ice and putting the tubes in a hot water bath for a heat shock for exactly 30 seconds. This was one of the most critical steps in the process because if the bacteria sat in the hot water bath for one second too long, the bacteria would simply die because of the heat and the whole process would be defective as the bacteria would be dead. We then moved on to plating the solutions in the test tubes to be placed in the incubator, in which we will then check back and hopefully see that the bacteria is glowing indicating the process was successful.

After we got our plates out of the incubator, we gathered the data that is presented below. Our specific experiment was somewhat successful in that we performed a successful transformation and ligation reactions which only allowed 2 colonies of bacteria to be present in tube #3. When we took 50 uL out of the tube to be put in the Amp+ plate, we just happened to bring the only 2 colonies that were produced into the plate which would not make the bacteria glow because of an absence of Arabinose. Because of this, there was no bacteria left for the 50 uL to place in the plate containing both Amp+ and Arabinose. As a result, we were left with 2 colonies in the Amp+ plate and no colonies in the ++ plate containing both Amp+ and Arabinose–which did not glow. This likely occurred because of the small amount of bacteria present following the transformation and ligation reactions.