By Senior Airman Ashlin Federick, Armed Forces Medical Examiner System / Published January 06, 2017
DOVER AIR FORCE BASE, Del. --
As I settled into my new assignment at the Armed Forces Medical Examiner System this past year, I embarked on a personal DNA journey. To be honest, I never really understood the in-depth specifics surrounding DNA and the potential influence it has in providing closure to military families.
Last year, I had the opportunity to attend a Defense POW/MIA Accounting Agency Family Member Update in Arlington, Virginia. This is an event like no other. Family members of those still unaccounted for from past conflicts gathered in one location to hear briefings from various experts and receive updates about their specific cases. I witnessed relatives of those lost contribute Family Reference Samples to the AFMES’ DNA identification lab. The reference samples can then be used as points of reference to compare the DNA of their unidentified loved ones to. This was all very interesting to me because I never really thought about DNA in this manner before.
Military members are required to have blood stained cards on file for the sole purpose of identification. A DNA database for military members does not exist. However, since I now work at AFMES and may go into the labs on occasion, it is necessary for my DNA sequence to be on file for exclusionary purposes in case of possible contamination. For this reason, I was able to go through the exact same process family members do by having my DNA collected and analyzed. I felt the need to follow this process and share my story. I have never been really good in science, so it took me a few minutes to learn all the scientific terms for what the analysts were doing, but after a day or so I began to understand.
The first day I walked into this bright, very clean laboratory, and on the table were some forms and these two black plastic sticks in a sealed bag. Jennie McMahon, AFMES Department of Defense DNA Registry supervisory DNA analyst, took the sticks, which I now know are called Bode buccal swabs, and proceeded to swab the inside of my mouth. They looked like large plastic Popsicle sticks, and it sure felt like someone was rubbing a dry Popsicle stick on my cheek as well. She did this two more times, then placed the swabs back into the bag and put them in a refrigerator.
A couple of weeks later I received an email that my buccal swabs were ready to be processed. Now, this is where things will get a little technical.
Jessica Hart, AFMES DOD DNA Registry quality control technician, and Marc Keirstead, AFMES DOD DNA Registry quality control analyst, showed me how they extract DNA from the sample. The machine looks like a giant hole-puncher that can punch two holes in the buccal swab at the same time and barcode it simultaneously when it drops into the plate. This was done for each of the samples.
After my DNA had been extracted, Hart went through the steps of amplification, product gel and purification with me.
Amplification is when they take your DNA and make copies of it so it can be read easier. This was done by making a chemical concoction and then transferring the extracted DNA into the mixture. The master mix and water is added to a new 96-well plate. A multi-channel pipette is then used to transfer the extracted DNA to the plate containing the master mix. The plate is then sealed and placed in a thermal cycler, which is programmed to cycle thought the specific temperatures and times required for amplification, for an hour and a half to two hours.
Once amplification is complete, my DNA was subjected to a method called gel electrophoresis. This was fascinating to watch because this process allows your DNA to actually be seen. Hart added the amplified DNA to a slice of gel as a way to see if the amplification worked. An electrical current was run though the gel for 15 minutes to separate the DNA by size. The dye then interacts with the DNA causing it to fluoresce and a photo was taken under ultraviolet light. This was to see the different sizes of the DNA. It was remarkable to watch because in the gel it just looked like a bunch of blue lines, but after the picture was taken with the UV light, you could see the various color differences.
Following the product gel, the amplified DNA was purified, breaking down any of the amplification reagents still in solution so they would not interfere with the sequencing.
Christopher May, AFMES DOD DNA Registry validation technician, took me into a laboratory where the sequencing is done. He set up a robotic liquid-handling machine with a bunch of tubes and then set plates into it. The machine transferred everything from the mixture as well as the purified DNA into multiple plates. After the plates were complete, another robotic machine, known as the rack runner, sealed the plates and put them in a thermal cycler.
After being removed from the thermal cycler, I watched May add chemicals to a machine called a DNA Analyzer, in order to bring the DNA back into a stabilized solution. It was then injected in the Capillary Electrophoresis instrument. This device uses a laser to excite the dye, which then emits a particular wavelength of light, red, yellow, blue or green, making it easy to identify DNA sequences based on the color produced. Blue corresponds with cytosine and green to thymine, two of the main bases that make up DNA.
I was offered the option to witness them analyze both my mitochondrial and nuclear DNA. For me, this was a once-in-a-lifetime experience, and I’m truly grateful I had the opportunity to be involved, while becoming more familiar with the methods and techniques used for processing and sequencing DNA.
During this process I learned that mitochondrial DNA (mDNA) is passed on through your mother’s side of the family, but nuclear DNA comes from both parents. This means if you were to put my mother’s mDNA and mine side-by-side, they would look exactly the same, but my nuclear DNA is unique to me, unless I had an identical twin.
Keirstead said there are differences between mDNA and nuclear DNA. Nuclear is inherited from both the mother and the father and is unique to an individual whereas mDNA is maternal and is common amongst anyone following the maternal line.
“As a result, nuclear DNA will provide you with more discriminatory power, but only if it is available,” said Keirstead. “In situations where remains are older or severely degraded, that may not be the case. Within a cell, you will only have two copies of nuclear DNA, where you would have more than 1000 copies of mDNA, making it an excellent tool and resource in identifying these types of remains based on the sheer number alone.”
From my mDNA, I found out that I belong to the haplogroup T2, which means my family more than likely descended from Hispanics. This came as a shock to my mother and grandmother who were unaware of any Hispanic lineage.
I was honored to be a part of this experience. It revealed to me the great lengths those who work within the DOD DNA Registry and AFMES are willing to go through to ensure past loved ones are accounted for in hopes this may provide families some closure they so deserve.
McMahon said she thinks the whole DNA process is amazing because each year, new advances are made allowing them to piece together more DNA puzzles using smaller portions of samples and ultimately identify more missing service members.
“I'm very proud to be associated with the Armed Forced DNA Identification Laboratory mission,” said McMahon. “I have been part of the past accounting section for 18 years and still get excited each time a sample gets reported out. I know that our work with DNA brings that individual one step closer to being identified and the family one step closer to having closure. We want each family to know their loved one has been found and will be returned home.”