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How to hide your DNA – the guide to leaving no traces

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A thorough guide to DNA, DNA transfer, and DNA destruction techniques

We’ve all watched our favorite detective show and screamed at the TV, “How could you be so stupid!” when the bad guy does something dumb and gets caught. We’ve thought to ourselves, I could have gotten away with that. But in modern times, I wonder if I really *could* get away with a crime given the prevalence of surveillance cameras on every street and doorstep, advanced satellites capable of tracking a person’s every movement, and, of course, the impossible task of leaving no DNA traces behind. Then I dug deeper (Dr. FBI Man, I have no intention of committing any crime; this is pure geek stuff, plain and simple) and found that with regards to DNA, it’s more than just wrapping yourself in a bubble or scrubbing a crime scene. But first, we must have a clear understanding of what DNA is, how it is collected and processed, and how it is used to identify a person beyond a shadow of a doubt.

About DNA and genetic markers

Every living organism, including humans, consists of cells, such as those in your skin. These cells contain sub-compartments that specialize in functions like energy production and molecule export. One key compartment is the nucleus, which safeguards the cell’s genetic material. This genetic information is encoded in a molecule known as DNA, or deoxyribonucleic acid. Another important compartment is the mitochondrion, which also contains some DNA. A human cell can have anywhere from 0 to 2000 mitochondria.

DNA holds unique genetic information known as the genome for each individual, apart from identical twins. We can imagine DNA as a book. Within this book, rather than coherent sentences, we find sequences of letters. Typically, these letter sequences are nonsensical, but occasionally, we come across recognizable words amidst them. For example, we can read:

JVLKHZKHEFLKAEJLKJFA**SMALLBROCHURE**CLZAEHDFILZAH

There are sections that have no apparent meaning, which correspond to DNA sequences that are called non-coding sequences. The segment *SMALLBROCHURE* in the middle is a piece that makes sense. This sequence could, for instance, be code for a trait in the cell. This is called a gene.

A gene refers to a segment of DNA that holds a unit of genetic information. The genome encompasses all genetic materials in a cell, including both coding and non-coding regions. Most of the human genome consists of non-coding sequences, which are portions that do not represent genes.

Genes can exist in different forms known as alleles. For instance, regarding an “eye color” gene, there are various alleles like “brown,” “blue,” “green,” and “gray.”

Structure of DNA

To reuse our book comparison, the information in the book is written using the 26 letters of the alphabet. DNA, on the other hand, is written using four letters, which are four molecular bases: A, T, C, and G.

492px Phosphate backbone

A DNA strand consists of four fundamental components known as nucleotides: A, T, C, and G. These nucleotides are arranged in a specific sequence, referred to as a DNA sequence. The length of a DNA segment is measured in base pairs (bp). When discussing a segment of DNA, we can denote its sequence, such as ATACCACAACATCACA. If this sequence encodes for cellular characteristics, it’s identified as coding DNA, while the remainder of the DNA is termed non-coding DNA.

DNA constructs a double helix consisting of two single strands linked by base pairing, where A pairs with T and C pairs with G. 

This book consists of multiple volumes, from 1 to 23. Each volume is available in “duplicates”; that is, there are two versions with the same layout (for instance, a specific word appears on page 3, fourth line) but differing content (the words may not be identical).

DNA animation

Each volume represents a chromosome, typically existing in pairs—one from the mother (maternal) and one from the father (paternal). For instance, the sex chromosomes are represented as XX or XY. The DNA double helix forms chromosomes by joining with other molecules. Humans possess 23 pairs of chromosomes, meaning every cell contains 23 pairs of these chromosome-books. Thus, we can describe an individual as the library in our example.

Stability of the DNA molecule

Let’s continue on the subject of DNA books. The content of these books is rarely modified over time because it is protected (by the cover, for example). To make this book unreadable, we try to cut it into small enough parts that we can no longer extract anything from it. For example, if we take the sequence we had before:

SMALLBROCHURE

And we cut it into many small pieces, we get the following: S, M, A, L, L, B, R, O, C, H, U, R, E. With just these letters, whether the base text was SMALL BROCHURE or MALL CHORE RUBS is impossible to know.

The book’s cover represents the proteins that encase and safeguard the DNA, alongside the nucleus, which protects everything. Attacking the DNA chemically is akin to cutting it, rendering it unreadable.

DNA is a very stable molecule, which means that it doesn’t change easily (whether from mutations or cutting the DNA strand). All this is to say that DNA is robust.

However, DNA can break down, resulting in the molecule being divided into increasingly smaller fragments. Because the information lies in the arrangement of the base nucleotides, shorter sequences provide less information.

Each cell contains just one copy of nuclear DNA, found in the nucleus, while mitochondrial DNA usually has several hundred copies. This abundance allows it to be analyzed even in older or more degraded samples.

How the police collect DNA

First of all, it is important to specify that DNA collection is not an automatic process: sufficient quantities and a degree of quality are required for it to be usable by the police.

Collection methods at the site

At the site of the action, the police will use these methods to take the DNA samples to the lab for analysis:

  • Liquid samples (liquid blood, spit): They are transferred to a sterile tube, refrigerated, and delivered to the lab as soon as possible. If the transport time is likely to be more than 24 hours, the sample is absorbed with a cotton swab or sterile pad and allowed to dry.
  • Solid residue (dried blood stains on clothing): If possible, the surface under the residue is cut out, bagged, and removed. If cutting isn’t possible, a sterile swab moistened with minimal sterile water is used to rub the sample. The cotton swab is then dried and preserved for analysis.
  • Touch DNA (skin cells inside a glove, cells inside a mask around of the mouth): The police rub the area with slightly damp cotton or a swab and store it in a sterile tube.
  • Solid bits (a hair, a piece of skin that is torn off): These are put in a sterile bag.

Usable DNA sources

Not all DNA that is collected is usable and the source of the DNA determines how effective it is for analysis.

Blood

Blood is the prime biological sample. All DNA in blood originates from white blood cells (5,000 to 10,000 cells per μL). Scientific analysis of blood is straightforward: theoretically, any visible trace can be examined.

Menstrual “blood”

Menstrual fluid consists of blood and discarded cells from the uterine lining. This is reportedly beneficial for genetic profiling.

Saliva

Saliva holds hundreds of cells per μL from the mouth. DNA from these cells can be found on licked items, like stamps, which is a well-known example. Even after months or years, a licked stamp remains suitable for analysis.

Urine

Urine is not an ideal source for DNA analysis due to its low DNA content and rapid deterioration.

Estrazione DNA cropped

Bones and teeth

DNA extracted from bones and teeth is primarily utilized for corpse identification. This genetic material has a high likelihood of surviving long after death, even when the body has undergone severe damage, such as burning.

Skin samples

Intact DNA profiles can be obtained from direct skin swabs.

Objects in contact with the skin

Examples include door handles, car keys, telephones, cup handles, knife handles, shoes, socks, clothing, glasses, hats, latex gloves, pencils, and watches. The success rates for “Touch DNA” analysis can vary significantly because the biological material left on surfaces differs greatly among individuals; for instance, dry, clean hands leave less DNA. Frequently, the DNA present is in very small amounts and may be degraded, resulting in only partial DNA profiles being obtained.

Keep in mind that smooth surfaces tend to retain fingerprints more readily, while rough surfaces are more prone to retaining DNA traces, as they can dislodge cells, for instance.

If you touch one object with a gloved hand and then touch another, you can transfer DNA from the first to the second. In general, contact between two objects allows for DNA transfer between them.

Hair and body hair

The success rate for DNA analysis from the nucleus of hair shaft cells is less than 10%. This is largely due to the minimal and highly degraded DNA found in hair. When hair falls out naturally, the root is similarly degraded, resulting in few or no viable cells.

Hair that has recently fallen out or has been pulled out retains its root, making nuclear DNA analysis easier. Mitochondrial DNA may serve as an alternative, though it is less effective than nuclear DNA for analysis. Additionally, non-human hairs can provide valuable information!

Cigarette butts

A classic scenario involves a friendly police officer offering you a cigarette while in custody. The likelihood of achieving accurate analysis in this situation is quite high. To ensure this, the paper around the filter and the final millimeter of the filter itself are meticulously removed, specifically the sections that come into direct contact with the lips and mucous membranes.

Glasses, containers, and food

Drinking from a glass leaves cells on the rim where your lips touch. This is known as “Touch DNA” and can be collected by law enforcement.

Stains on fabric

If a stain dries before the fabric is washed, there’s a high likelihood of residual DNA after washing. For blood stains, even after a machine wash at 90°C, they remain analyzable. Similarly, stains can still be examined after hand washing with soap, dishwashing liquid, or laundry detergent.

How DNA identifies a person

Genetic profiling relies on the fact that only twins possess identical genomes. While the human genome is largely uniform across individuals, variations exist that can be utilized to identify a person, known as genetic markers.

Genetic markers used by the police

There are different types of variations between individuals. The ones used by the police for identification purposes are called **STRs** (Short Tandem Repeats), also called microsatellites.

What is an STR (Short Tandem Repeat)?

Let’s return to our DNA-book comparison and take a closer look at a sequence:

JVLKHZKHEFLFA**HLHLHLHLHLHLHL**AEJLKJSMALLBROCHUREFILZAH

Here’s a sequence in which the pattern *HL* is repeated seven times. The same thing can be found in DNA, for example, with repeats of the CA pattern. The Short Term Repeats in the above example would be the entire sequence HLHLHLHLHLHLHL, consisting of 7 repeats of the HL pattern.

In the example above, the STR is located in an area of the DNA that does not make sense (non-coding DNA), but it could also be located in the middle of a part that does make sense (coding DNA). For example, below, the STR is located right in the middle of SMALLBROCHURE:

JVLKHZKHEFLAEJLKJFASMALLBROHL**HLHLHLHLHLHLHL**CHUREFILZAH

The STRs analyzed by the police are located in the non-coding regions.

The different alleles of a STR

Let’s imagine that in all libraries (i.e., in all individuals), in book 4, page 32, 4th line, there is always a certain number of repetitions of the HL pattern. In some libraries, at this precise place, there are 8 repetitions of HL, while in others, there are 7, 4, or 12, for example.

In individuals, it is the same. Let’s imagine a particular STR made up of repeats of the pattern GTC, located on the 3rd chromosome, starting at 2 million base pairs from the end of the chromosome. Some individuals will have 8 repeats of the GTC pattern, while others will have 12, or 7 or whatever.

The count of repeats in a specific STR can vary. In the general population, there can be up to ten distinct versions of a particular STR, with each allele defined by its unique number of repeats. In this context, we adopt a broader definition of allele: for a STR, the allele refers to the number of repetitions of the repeated sequence.

Genetic profiling determines the alleles of a given number of different STRs. For example, by studying different STRs numbered from 1 to 5, the number of repeats of the pattern (i.e., the allele) for each STR can be written as:

16/15(STR1); 15/7(STR2); 13/12(STR3); 5/6(STR4); 8/8(STR5)

For STR1, there are 16 repeats on one chromosome and 15 repeats on the other.

Each STR can have two different repetition counts due to the presence of two chromosome versions: “maternal” and “paternal.”

Why STRs are good genetic markers

Forensic requirements for good genetic markers are as follows:

  • They must be stable over a lifetime. STRs are carried by DNA, a fairly stable molecule over a lifetime.
  • They must be resistant to the degradation that organic structures may undergo. STRs are rather short (< 300 bp), so they have little risk of being affected by degradation. In essence, selecting a shorter DNA sequence reduces the likelihood of degradation compared to longer sequences.
  • The analysis technique required must be sensitive enough to apply to minute traces. This sensitivity is what has improved the most in the last 20 years.
  • The cost of the analysis must not be prohibitive.
  • There must be enough polymorphism, i.e., enough variation in the population, for the marker to be useful for identification. STRs have many alleles, i.e., many variants.

DNA analysis by forensic police

How is DNA analyzed? The sample is first purified, meaning it is cleaned to keep only the DNA (all other substances are removed).

Next, the collected DNA undergoes amplification, meaning it’s copied multiple times. This process utilizes PCR, or Polymerase Chain Reaction, a technique for molecular copying also referred to as gene amplification. In PCR, a specific segment of DNA is chosen and replicated millions of times.

After PCR, the products are examined by gel electrophoresis. Electrophoresis is a method used to separate molecules based on their varying mobility in an electric field. For example, the more positive charges a molecule has, the more it will be attracted to the negative pole in an electric field, so the more it will move.

In gel electrophoresis, the molecules move in a gel: the larger they are, the less they move because the gel hinders their movement.

There are, therefore, two parameters that allow molecules to be separated in gel electrophoresis: their charge and their size. Since gel electrophoresis allows separation according to the size of the molecules, it can, therefore, determine the size of a piece of DNA by comparing its migration with the migration of pieces of DNA of a known size.

This technique is used for several STRs (multiplex analysis). This provides a sample’s DNA profile. Another widely used test is to determine whether the sample contains a Y chromosome.

The DNA profile that is obtained corresponds to the number of pattern repeats in a given number of STRs (15 STRs for the analysis kit called ABI used in France, for example).

What influences the success of collecting the genetic material from the site?

The amount of DNA found at the site

The recommended number of cycles in PCR is 30. This allows a satisfactory DNA profile to be obtained from 0.5ng of DNA. Considering that we have about 0.006ng of DNA per cell, it takes the genetic material of about 100 cells to obtain a profile (in 2006).

We will use these numbers for our calculations, but the amount of DNA needed for analysis is decreasing all the time.

There are different amounts of DNA in different sources:

  • In one mL of blood, there is about 30,000 ng of DNA. It takes at least 20 millionths of a mL to do a DNA analysis that gives a satisfactory result. This is smaller than a small drop of water.
  • A sample taken directly from the skin of the hand yields between 2 ng and 150 ng of DNA (on average 50 ng). Dry or recently washed skin will yield less DNA than other skin.
  • One mL of saliva contains about 2,000 ng of DNA. Therefore, it takes at least about one-millionth of an mL to do a satisfactory DNA analysis.
  • One hair root yields up to 200 ng of DNA.
  • In one mL of urine, 10 ng of DNA can be found.
  • One mg of skin or muscle contains about 3,000 ng of DNA.
  • A swab from objects held regularly by an individual yields between 1 ng and 100 ng of DNA.

When police have only a few cells available, the analysis is much more vulnerable to random effects that do not reflect reality. For example, consider 3 cells containing the A and B alleles of a given gene. When these cells undergo a degradation, the B allele does not survive. Only the A allele remains each time. The police collect a sample and analyze it. They obtain a profile where only the A allele appears, so their interpretation will necessarily be incomplete. This is called an allele drop-out.

Even if we always leave some DNA, it is worthwhile to leave as little as possible. It is not a question of all or nothing.

DNA preservation and degradation

DNA can be degraded before or after it is collected if it is poorly preserved by the police who collect it.

To successfully analyze a DNA sample, researchers require several DNA fragments that are a few hundred base pairs long (nucleotides). Nonetheless, various environmental factors can lead to DNA degradation (break it).

  • Certain enzymes found in the environment, such as those produced by bacteria or fungi in soil, can cleave DNA. They operate optimally at specific temperatures.
  • Humidity supplies one crucial ingredient—water—essential for breaking the bonds between nucleotides in the DNA chain. Water is required for the action of DNA degradation enzymes. Additionally, water fosters the growth of bacteria and fungi, which recycle fragments of the sample’s molecules to create their own molecules, including DNA.
  • Heat is one of the elements that accelerate chemical reactions that require energy, which include DNA degradation reactions.
  • Radiation can damage DNA by causing strand breaks or forming pyrimidine dimers.
  • Similarly, light (especially UV light) chemically transforms DNA. Some skin cancers are linked to UV, for example, because UV tends to mutate DNA.
  • Other aggressive chemical substances (acids, bleach) can also attack DNA molecules. However, these substances are unlikely to be encountered at crime scenes. We’ll expand on that a bit later (the benefits of leaving chemical substances at the site).
  • Outdoors, the sun, and rain leave little hope for biological traces to last beyond a few weeks to a few months. If the area where the DNA sample is left has significant microbial life (e.g., undergrowth, soil, stagnant water), this lifespan decreases further. A hammer left in the undergrowth in the middle of summer (enzymes, humidity, heat) will keep less DNA than if it was left on concrete in winter.
  • Moreover, to take saliva as an example, when it is not dried, the humidity and the enzymes present rapidly degrade the DNA (in a matter of hours or days).

To be properly stored, DNA must be dry, cold and protected from light. The recommended storage conditions for forensic samples are room temperature and low humidity for dry samples, and -20 °C for liquid samples.

Analysis of samples containing DNA from multiple individuals

Analyzing samples containing multiple individuals’ DNA is a significant challenge for police DNA forensics. It is also a field where lots of research occurs, and progress is quick.

Two important factors influence the success of an analysis of mixed DNA:

  1. How many individuals contributed their DNA to the mixture? (the more people, the more difficult to analyze)
  2. How much did each individual contribute to the mixture? For example, if one person contributed a lot, and the others just left a very small amount of DNA, it will be easier to analyze than if everyone contributed about the same amount. Also, the smaller the amount left by each person, the harder it is to analyze.

How degraded is the DNA?

This area has advanced a lot, with new standards and statistical tools to address it. There are several steps to analyze a DNA mixture:

  1. First, identify that it is a mixture.
  2. Find the peaks of alleles (like on the graphs).
  3. Deduce the number of potential individuals whose DNA is included.
  4. Find the contribution ratio of each individual to the mixture.
  5. Find all the possible genome combinations.
  6. Compare with DNA profile databases.
  7. Pray you didn’t screw up (apparently, this is important).

Location of the collected DNA

The site from which DNA has nothing to do with the quality of the analysis. Instead, it influences the interpretations that police or judges may make. The connection between the sample’s origin and its collection location enhances the usefulness of DNA evidence for incrimination.

Blood, whether discovered near or at the scene, serves as strong evidence. In contrast, a cigarette butt showing your DNA found 50 yards from a ransacked bank holds less weight than your blood at the same spot. Thus, the location of DNA sources other than blood makes a substantial difference regarding their incrimination. Transportable (or mobile) sources of DNA (e.g., hair blown in the wind, bits of skin left on an abandoned bag in the street) are weaker evidence than others (like blood).

Thus, it’s important to keep in mind that just because the police collected your DNA doesn’t mean you’re screwed. What they want most of all is a confession, and DNA may very well be all they need to get it.

The genetic fingerprint kept by the police

Once a saliva sample is taken by law enforcement (a cotton swab inside your cheek), it is packaged and sent to a laboratory for analysis. Once the analysis is finished, the extracted DNA profile is presented as a table.

What the police are recording in that table is a sequence of numbers that they hope is unique to each individual. By refusing to give a DNA sample, you refuse to improve the police’s knowledge of our genetic information. Every DNA sample taken by the police is a measurement of tiny parts of the genetic information contained in your DNA and not a complete picture. Even if you’ve already been swabbed, giving another swab means the police are storing new information about your DNA. When in doubt, it’s always best to refuse a sample, even if you think your DNA has already been collected somehow.

How to avoid leaving DNA (or destroy it if it’s been left behind)

The judicial system often utilizes DNA evidence, but misinterpretations can lead to innocent people being imprisoned. Our aim is to clarify how law enforcement identifies individuals through DNA and to suggest methods for safeguarding against potential identification . . .

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Additional notes

Note: Uses (or not) of STRs

There are many different kinds of STRs, with different “uses”.

For example, there may be STRs in the actual coding parts of genes, which will result in repeats of the same amino acid in the protein produced from the gene. STRs can also appear in the regulatory sequences of genes (the sequences that say “please express this gene a little more” or “not this one”). The roles we are discussing here are not well understood, to say the least.

There are also STRs at the end of chromosomes, because every time DNA is copied (for cell division for example), the chromosomes are not copied to the very end. This is one of the reasons why we age. The STRs at the end therefore serve as buffers that can be shortened at each DNA replication without risking damage to the genes. In short, they prevent our genes from being nibbled at during each cell division.

But here we have talked about the evolutionary uses of STRs, but there are also STRs present that do not contribute anything to the organism that carries them. In fact, at the molecular level, mutations are mostly random and are not selected for their evolutionary value. This is the neutralist theory of evolution that was put forward by Motoo Kimura.

Note: Techniques used on DNA during analysis

Prior to analysis, DNA from a sample is isolated and purified. In forensic methods, DNA probe techniques are used to pre-test the presence of human DNA in a sample before proceeding with the actual analysis.

PCR (Polymerase Chain Reaction)

PCR is a molecular copying technique. It can be called amplification. During PCR, not all the DNA present in a cell is copied: the copying selects for a small segment of DNA using the primers, which are positioned at both ends of the area to be copied and thus delimit the copying range.

PCR Process

The process begins with a piece of DNA that contains the part to be amplified. It is combined with a large excess of DNA pieces that are a few nucleotides long (called primers). These primers are complementary to a precise region of the DNA that they want to amplify. DNA replication enzymes (DNA polymerase) and nucleotides, necessary for the synthesis of a new DNA strand, are also added.

At 94°C, the hydrogen bridges between the two DNA strands are broken (the DNA is denatured).

The temperature is lowered to between 50°C and 60°C. At this temperature, the two strands could reassemble (the hydrogen bonds can be reformed). However, there is a large excess of primers compared to the two initial DNA strands. Most of the hydrogen bonds that will be reformed will therefore be between the initial DNA strand and the primer, which is the black strand with an arrow on the following diagram. Here, we have only made the diagram of one of the two initial DNA strands: exactly the same thing happens on the other strand!

The temperature is increased to 74°C, which is the optimum temperature for the DNA polymerase. This enzyme will then replicate the DNA by attaching itself to the primers to start its synthesis. The black circle represents the DNA polymerase.

This cycle can be repeated for as long as they like. Starting from one DNA molecule (double stranded), they obtain 2 double stranded DNA after one cycle, 4 after 2 cycles, etc. So they have 2 to the power of n double stranded DNA molecules after n cycles. The number of cycles is normally 28.

Multiplexed STR analysis

PCR is easy to use and can amplify several DNA fragments simultaneously (just add primer pairs). This is called multiplex PCR.

Variants of PCR used

Several types of PCR are used, each with advantages and disadvantages.

Low Copy Number (LCN) is a DNA profiling method that uses a larger number of cycles for PCR (34 instead of the traditional 28), which makes it both more sensitive to smaller amounts of DNA, and more vulnerable to contamination, among other disadvantages.

Direct PCR is the process of performing PCR without isolation or prior purification of the DNA sample. There is less risk of losing the little DNA present in the sample and less risk of contamination (because there is no isolation or purification), but the subsequent analysis is more time consuming (because there is more chance of ending up with messy mixtures of DNA).

Note: Principles of DNA sequencing

The first sequencing technique invented was the Sanger technique. It uses the principle of chain terminators. A chain terminator is a nucleotide that can bind to a nucleotide during replication, but to which the next nucleotide cannot bind.

It starts with the molecule to be sequenced, a large batch of nucleotides, and chain terminator A nucleotides (i.e. nothing can attach after them). So there is a very large number of copies of the DNA strand they are interested in, and it will be copied with enzymes. At each position where an A must be integrated, some molecules of this very large number of molecules will receive an A terminator. Their copy stops there. In the end, we get a lot of partial copies, with an A at the end. By finding their length, we can deduce all the positions of the A’s in this sequence. We can then do the same thing with C, G and T!

There was a time when sequence polymorphism was used for forensic DNA analysis, i.e., whether the DNA used had the *ATTACG* sequence or the *ATTACC* sequence. Now they use length polymorphism, by analyzing the lengths of repetitive sequences, i.e. they try to know if *ACG* for example is repeated 10 or 12 times in the sample.

Capillary electrophoresis and multiplex analysis

Today, there is also the capillary electrophoresis technique, which allows for much larger potential differences and therefore faster migrations. It is the same principle as gel electrophoresis, but instead of being done on a gel plate, it is done in a capillary. It is a state-of-the-art tool used by forensic police.

Limitations of multiplex analysis and partial solutions

Having several STRs poses a problem for electrophoresis: each individual has a maximum of 2 different alleles for each STR, so if they analyze 2 STRs at the same time, they have 4 potential alleles. The challenge is to know to which STR each allele belongs to. This can be solved by taking STRs of different lengths. For example, if, for a gene, all the alleles are between 100 and 170 bp, and for another gene, all the alleles are between 200 and 250 bp, they can be analyzed at the same time by electrophoresis without risking confusion.

However, they cannot analyze as many STRs as they want all at once, because this implies having larger and larger STR alleles, and the effectiveness of PCR has size constraints. So there are limits to simultaneous STR analysis.

Another solution is to attach fluorescent primers on the different STRs to differentiate them at the time of electrophoresis analysis.

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