Crime Library: Criminal Minds and Methods

All about the DNA Revolution

How DNA Analysis Works

Since 1985, the use of DNA fingerprinting, better known among professionals today as genetic identification, DNA profiling or DNA analysis, has taken one of two basic directions, and one type of testing needs very little evidentiary material for linking a specimen with a probable originator. The first procedure in the discovery of DNA identification is called Restrictive Fragment Length Polymorphisms (RFLP) analysis and the second is Polymerase Chain Reaction, or PCR. Both are explained below.

DNA expert Keith Inman has been a criminalist for the Sheriff's Department in both Orange and Los Angeles Counties and for the Los Angeles County Chief Medical Examiner. He also worked at Forensic Science Services of CA, Inc, a private crime laboratory, and is currently employed by the California Department of Justice DNA laboratory. With Dr. Norah Rudin, Inman has written An Introduction to Forensic DNA Analysis. Despite the popular conception that DNA profiling is similar to fingerprint technology, he explains that the term 'DNA fingerprinting' is a misnomer: "While real fingerprints can distinguish between identical twins, DNA analysis cannot." Comparing it to fingerprinting raises misguided expectations.

"DNA profiling," says Inman, "examines small sections of human DNA that are known to vary among people. It is similar to ABO blood group typing in the sense that different people have different types, but is far more discriminating. DNA has the potential to discriminate between all people, except for identical twins."

Early in the century, Phoebus Levine made the discovery that individual cells contain two types of acid in the nucleus: ribonucleic acid (RNA) and deoxyribonucleic acid (DNA). Within each cell's nucleus are twenty-three pairs of chromosomes that are made of DNA, which acts as a blueprint to dictate our physical functions and characteristics. The "instructions" are issued in a genetic code transferred from our parents and these are unique to each person. To understand how the analysis works, we must first understand the make-up of DNA.

Each DNA molecule contains four chemicals units:

Adenine (A)
Guanine (G)
Cytosine (C)
Thymine (T)

When strung together in paired chromosomal strands (double-stranded helix), A always aligns with T, G with C, etc., to form the protein and enzyme make-up of our cells. In structure, they have the appearance of a twisted ladder, with these alternating pairs as the rungs. Inside the cell, DNA is tightly coiled, but when unrolled, a molecule of DNA is approximately six feet long. Although some parts of our DNA are universally human (species-specific), certain sections contain the codes that give us our individual uniqueness, and this DNA alignment is the same in every cell in that person's body. These variations in base sequence are called polymorphisms because they vary in shape from person to person. Thus, by looking at the parts of the DNA that make a person unique, experts can determine whether a particular strand of DNA found in a specimen is indistinguishable from the DNA of a particular person.

The polymorphic DNA regions repeat themselves over and over. Some inform our development and the function of others is unclear. The base pairs in these regions are called Variable Number of Tandem Repeats, or VNTRs, and they provide the possibility for genetic identification. To find the polymorphisms, one must first separate out the DNA from any protein that is attached to it.

"DNA must first be removed from the surrounding cellular and environmental materials," Inman explains. "This is called extraction." What happens next depends on which test is used.

In RFLP testing, the extracted DNA is mixed with a substance called a "restriction enzyme" that "digests" or cuts the DNA strand in different areas. Those fragments are then covered in a gel to separate the double-sided fragments into single strands and electrical current is applied (electrophoresis). The negatively-charged fragments move through the gel at different speeds toward the positive pole, with the shorter pieces migrating faster. The end result is that they line up according to size.

The scientist then takes the pieces from the gel with a nylon membrane called a Southern Blot, and the DNA fragments get fixed to the membrane through heat or air-drying. The A, T, C, and G bases of the strand become exposed and ready. They get treated with a radioactive synthetic genetic probe—a DNA fragment that acts as a sort of sleuth. The single-strand probe seeks out and binds to its complementary base, revealing a pattern. (The multi locus probe would bind to multiple points on multiple chromosomes, while the single locus probe focuses on a region within one chromosome.)

The probe identifies some pieces of the DNA with dark bands, as revealed by an X-ray (autoradiograph or autorad) of the membrane. This looks similar to a bar code found on food at the supermarket. Then a print is made of the polymorphic sequences, which can be compared (visually and by computer) to prints made from other specimens. For example, when a woman was killed, the saliva left behind on a bitemark was compared by DNA analysis to a blood sample from the suspect to determine if that person had been present at the crime scene. (In a more recent development, a chemical luminescence is used instead of radiation, and it takes only a few hours to light up the pattern rather than days or weeks.)

The interpretation of a sample is based in statistical probability. If four fragments are identified, then the probability of each occurring in the population is multiplied against that of the other samples.

Criminal investigations often rely on blood samples from the accused and from the victim, but they may also use other biological products such as hair follicles, saliva, semen, tissue, or urine. The estimate of any two people having the same DNA can be as high as one in several billion.

"The common question being answered," Inman points out, "is, 'Who is the source of this biological material?' DNA profiling has had the greatest impact in the area of sexual assault evidence examinations. Prior to DNA testing, laboratories were very limited in the amount of genetic information that could be developed from evidence containing semen. With the advent of DNA testing, the genetic content of the sperm itself can be examined."

The PCR method has been hailed as an even greater breakthrough, although it is less precise. Whereas the RFLP method needs a considerable amount of material for testing (a semen stain the size of a nickel, for example) and may be unable to determine anything from a degraded sample, PCR theoretically needs a minute amount, because it works by mimicking the cell's ability to replicate DNA. It can also work with degraded specimens.

Also known as molecular xeroxing, PCR begins with extracting the DNA. Then it is heated in a thermocycler to make it split. The temperature is lowered and then raised again, which produces a copy (amplification). The process is then repeated.

"Chemicals are added," Inman explains, "that will locate the specific regions of the DNA of interest to us, and millions of copies of this DNA are made. Finally, the so-called amplified DNA is subjected to a typing protocol that identifies the specific types present in that region of DNA from the particular individual."

From identifying remains to deciding paternity to convicting criminal suspects, DNA testing has become a powerful tool in forensic services. However, there are some misconceptions.

"DNA provides powerful identification information," says Inman, "but the relevance of the evidence itself is always a case-specific question. With that in mind, we must realize that DNA doesn't really solve cases. It simply provides evidence that must be examined in light of all of the other evidence in a case. Sometimes, the DNA results can be highly inculpatory, yet at others be meaningless. Finding a semen stain on a bedspread in the local Rent-By-The-Hour motel does not mean much, even if you know from DNA testing from whom it originates."

Even so, this procedure is constantly being refined and will no doubt affect the future of criminal investigation, from aiding in conviction or in exoneration. Let's look at another case that made DNA testing into a national human-interest story. At the time, the results weren't known, and the story's climax surprised a lot of people.