Frequently Asked Questions

Maternal family members are eligible to provide blood samples for the purpose of establishing a comprehensive family reference DNA database.
A family reference database that includes an mtDNA sample for each of the unaccounted-for servicemen enables comparisons between remains in support of the identification process.

DNA (deoxyribonucleic acid) is a double stranded molecule of helical structure containing the genetic code that makes you who you are. You inherit your DNA from your parents and it determines certain characteristics such as hair color, eye color and other physical attributes..

 All DNA is constructed of the same four nucleotide bases: Adenine (A), Guanine (G), Cytosine (C) and Thymine (T), which are organized into two complementary helical strands. The number of Adenine bases is equal to the number of Thymine bases while the number of Guanine bases is equal to the number of Cytosine bases (A=T and G=C). For all DNA, a base-pair is considered an “Adenine base” pairing with a “Thymine base” on the opposite strand or a “Guanine base” pairing with a “Cytosine base” on the opposite strand. It may be easier to think of double stranded DNA as being arranged like a ladder where the sides of the ladder represent the individual strands of the DNA molecule and the rungs of the ladder are comprised of A-T or G-C base-pairs. The rungs of the ladder can contain either nucleotide base of the pair, and either base could be on either strand. 

 Autosomal and Y-chromosomal DNA are found within the nucleus of each cell of the body. There is a single copy of nuclear DNA, which is comprised of autosomal and Y- chromosomal DNA. Mitochondrial DNA (mtDNA) is found in the mitochondria of the cell. Mitochondria, found in the cytoplasm, or “body,” of the cell, are like batteries or powerhouses—they provide energy to the cell. There are hundreds to thousands of mitochondria per cell. Each mitochondrion contains its own DNA, separate from the nucleus. Even after many years, during which time all DNA degrades to some extent, mtDNA can be found in very small fragments of biological material. If there is sufficient quality, it can be tested and a sequence can be generated.

 Autosomal Short Tandem Repeat (auSTR) and Y chromosomal STR (Y-STR) tests are used to analyze nuclear DNA, Mitochondrial DNA control region or whole genome sequencing is used to analyze mitochondrial DNA. All thee DNA tests can be used to aid in the identification of missing service members. Because of the age and degradation of the DNA due to environmental conditions, mitochondrial DNA testing is the most sensitive and is usually the first type of DNA testing used. If the appropriate reference materials are available, autosomal DNA and Y-DNA will be tested as well. All of the DNA information can be used to calculate a combined likelihood statistic. The likelihood statistic assesses the evidential support for the identification hypothesis that the DNA from the unknown sample is biologically related to the associated references (auSTR, Y-STR and mtDNA).

 Autosomal Short Tandem Repeat (auSTR) and Y chromosomal STR (Y-STR) tests are used to analyze nuclear DNA, Mitochondrial DNA control region or whole genome sequencing is used to analyze mitochondrial DNA. All thee DNA tests can be used to aid in the identification of missing service members. Because of the age and degradation of the DNA due to environmental conditions, mitochondrial DNA testing is the most sensitive and is usually the first type of DNA testing used. If the appropriate reference materials are available, autosomal DNA and Y-DNA will be tested as well. All of the DNA information can be used to calculate a combined likelihood statistic. The likelihood statistic assesses the evidential support for the identification hypothesis that the DNA from the unknown sample is biologically related to the associated references (auSTR, Y-STR and mtDNA).

 The DPAA forensic anthropologists are very good at determining whether a skeletal element is of human origin by examining various bone features. However, some highly fragmented skeletal material may not be anthropologically distinguishable as human and may require DNA testing to confirm whether DNA obtained from the skeletal element is from a human or an animal. In 2012, AFMES-AFDIL developed a DNA procedure which amplifies a specific region of the 12S ribosomal RNA located within the mitochondrial genome. This region is species specific, which allows AFMES-AFDIL to determine if the DNA is human or non-human.

 To understand where the science is going we need to understand where the science has been and the continued advancements that have occurred over the past 20 years. The modern DNA forensic age started in the late 1980’s with the development of the Polymerase Chain Reaction (PCR), which allowed for the amplification of small amounts of DNA. In the 1990s, the first commercial 4 auSTR locus kits were released and AFMES-AFDIL developed the first mtDNA primer-sets; a commercial 7 auSTR locus kit was released in 1997 and AFMES-AFDIL simultaneously developed and published the first mtDNA mini-primer sets for processing degraded samples; a commercial 16 auSTR locus kit was released in 2001; commercial mini-STRs and Y-STR kits were released in 2005; commercial 16 auSTR locus kits optimized for inhibition were released in 2010; and commercial 20 locus kits were released in 2013. Each new commercially-developed kit brought greater discriminatory power, as well as increased chances of recovering usable DNA results from challenged samples. However, these commercial kits were optimized for criminal casework samples, which have high levels of DNA present and not the highly degraded skeletal elements processed by AFMES-AFDIL in support of the past accounting mission. During all of these changes, AFMES-AFDIL has continuously either optimized existing methods or kits, or developed new methods for processing challenged samples. The forensic DNA field is constantly evolving. AFMES-AFDIL stays abreast of all new technologies and leads the DNA forensic field in implementing or developing these state of the art technologies. For example, in 2015, AFMES-AFDIL discussed Next Generation or Massively Parallel Sequencing instruments which can sequence your entire genome in 48 hours. This technology is no longer the future, but is currently being employed by AFMES-AFDIL to assist with the past accounting mission (refer to Does AFMES-AFDIL have a Next Generation Sequencing Method?) However, building on the functionality of NGS instrumentation, AFMES-AFDIL is currently researching Identity Single Nucleotide Polymorphisms (i-SNP) to develop an identification panel. This is similar to what commercial entities are using for developing your personal genetic profile but more definitive.

 DNA is an integral part of the identification effort. This is particularly true when the skeletal elements are highly fragmented or commingled. DNA testing allows DPAA anthropologists to sort recovered elements into discrete groups, allowing for more efficient anthropological analysis. In other cases, where the skeleton has been broken into small fragments, anthropology may be limited in what it can tell about the samples and DNA will be the primary method of identification.

 In some instances, DNA analysis can provide a means of scientific identification, or definitive proof of your loved one’s identity. For example, if alternative reference materials (i.e. eye glasses, envelopes, service covers) are available from the missing service member, nuclear DNA may be able to be recovered from those materials and tested with an auSTR kit to provide a direct auSTR reference for the missing service member. When and if an auSTR result is generated from the skeletal samples recovered by the DPAA, the auSTR DNA profiles from the direct reference and the skeletal sample can be compared. If auSTR DNA profiles match exactly at a statistically significant number of loci, this will result in a positive identification. This scenario is what happens with current day military identifications. However, in most instances, the DNA reference materials on hand are from relatives of the missing person. This provides what is known as a presumptive identification. Regardless of the type of DNA testing method (auSTR, Y-STR or mtDNA sequencing), there is a statistical likelihood that the samples tested belong to your missing service member. When applicable, scientists at the AFMES-AFDIL will combine all of the available DNA testing information and calculate a combined likelihood statistic. The likelihood statistic assesses the probability that the DNA from the unknown sample is biologically related to the associated references (auSTR, and/or Y-STR and/or mtDNA profile)

If you are a maternal relative (maternal mother, maternal aunt, brother, sister [mtDNA testing]), a paternal relative (father, brother, paternal uncle, paternal cousin [Y-STR testing]), or a nuclear relative (father, mother, brother or sister [auSTR testing]) of a missing service member, you are eligible to donate a DNA reference material. Depending on your relationship to the missing service member you could be a reference for multiple testing methods (mtDNA, auSTR and/or Y-STR). For example, if the father is the missing service member and is survived by his son, his son is an eligible nuclear (auSTR) and paternal (Y-STR) reference. AFMES-AFDIL is attempting to collect at least 2 mtDNA references, 2 Y-STR and 2 auSTR references for each missing service member.

 As of June 2018, the percent of missing service members that have some type (mtDNA, auSTR, or Y-STR) of a family reference on file varies by the conflict:
• Vietnam War – 85%
• Korean War – 92%
• Cold War – 85%
• WWII –6%

Efforts for family reference collection are ongoing. If you or someone you know is a valid DNA reference donor for a missing service member please contact the respective service casualty office for information on participating. For World War II collections, family references are being collected on an as needed base due to the fact that a large number of missing individuals were lost in deep water losses and are not recoverable at this time.

 Forpresent-dayy accounting, all samples are compared to a direct blood reference to the deceased individual. Due to the success of DNA testing on past accounting samples, the United States Government, in 1992, established the Armed Forces Repository of Specimen Samples (AFRSSIR) to house and protect DNA blood reference cards for all active duty military, Reserve and National Guard personnel. The references and samples associated with present day accounting contain high amounts of DNA and only require single auSTR amplification for the reference and sample for a DNA comparison report to be issued. This differs from the past accounting mission, in which direct references are not an option in most cases and maternal, paternal and nuclear family references are needed to aid in the identification process. In addition, due to the low quality of the skeletal samples submitted by the DPAA for testing, all mtDNA, auSTR and Y-STR analyses must be performed at a minimum in duplicate in order to make a comparison. Duplication of the results is essential whether testing mtDNA or nuclear DNA as it is very easy to amplify a modern contaminant over the low quality authentic DNA.