Certagen GmbH

Parentage verification using DNA

Complexity of genetic material

The body of an organism consists of a large number of cells, which contain a full and complete set of genetic material. Genetic information is present in the nucleus of a cell. The genetic information is stored in chromosomes, which are translated by the body in useful data (proteins). This happens constantly in all cells. The general code is called DNA.

Chromosomes exist of long DNA-strains which are wound around each other very tightly. When a chromosome is studied in detail, it is possible to look at the composition of DNA in the form of A, T, G, or C. These A, T, G and C are the building blocks from which the DNA is constructed. Sometimes stretches of repeats are present (e.g. CACACA) – such stretches are indicated as microsatellites (also known as STRs). Other variation such as G/A or C/G is indicated as Single Nucleotide Polymorphism (SNP). The order and composition of DNA are the basis for all kinds of applications.

For the typing of the composition of genetic characteristics it is possible to use hairs, feathers – to be drawn with roots –, blood, milk, tissue etcetera. The usability of sample material depends on the test which is carried out. The use of ‘fresh’ material provides the best result.

 

Techniques

Genetic variation can be visualized with a number of different techniques. Frequently a technique is used, where DNA is multiplied (PCR). DNA can be made visible through three steps:

  • DNA-extraction, where the cells are broken into small pieces. The DNA is present in an aqueous solution, which is necessary to enable a successful PCR-reaction,
  • Selective multiplying of DNA, where PCR is used to multiply small specific pieces,
  • Analysis of DNA on a machine, with which DNA is visualized. For that purpose, fluorescence is incorporated during PCR.

The final result of these steps results generally in the detection of the variation in STRs or SNPs. By examining a number of STRs or SNPs, a genetic bar code is generated. This bar code can be used for a number of different tests, among which are lineage, identity of samples, etcetera. In a number of examples these applications are described below.

Parentage verification using microsatellites (STRs)

The genetic variation which is present in an animal, originates from both parents. Half of the variation is originating from the father, whereas the other half comes from the mother.

For parentage verification, typically 20 up to 40 genetic characteristics are visualized. In this process the length of genetic fragments is being measured. The measured length of a genetic characteristic in an offspring must correspond to the length in the mother and father that were provided for comparison. In two examples, it is shown how the basic rules are applied in parentage verification.

In the figure an example is provided of a correct parentage. In this figure, the DNA is shown of three individuals: an offspring (upper line), a potential mother (middle line), and a potential father (bottom line). In each line one genetic marker is shown. Two DNA fragments are visible as peaks. The first fragment of the offspring is originating from the father (length of the fragment is 150), whereas the second fragment comes from the mother (fragment length 152). In this case both fragments of the offspring are present in the parents: the parentage is correct.

In the second example a situation is shown where parentage does not qualify. The three lines are shown in the order of offspring, potential mother and potential father. Again in each line one DNA marker is shown, where two DNA fragments are visible as peaks. The second fragment of the offspring is present in the mother (fragment length 152), whereas the first fragment in the offspring (fragment length 150) is NOT present at the assigned father. In this case, one fragment is present at the offspring, which is not present in either of the parents: the parentage does not qualify.

When 20 up to 40 different genetic fragments are checked, the chance that an incorrect parentage is not detected becomes very small. The genetic fragments which are used for parentage verification and identification provide no information on properties such as color and quality of an animal, plant or human, since the fragments are non-coding.

 When the length of a number of DNA fragments is measured for a sample, a DNA-profile is established. This pattern is unique for a specific individual person, animal or plant, so that in cases of doubt DNA-profiles can be compared to confirm if two samples originate from the same individual.

Parentage verification using Single Nucleotide Polymorphisms (SNPs)

As with STRs, the basis is the same. However, more genetic markers are tested because the information content per markers is lower for SNPs.

The genetic variation which is present in an animal, originates from both parents. Half of the variation is originating from the father, whereas the other half comes from the mother.

For parentage verification, typically 200 up to 400 genetic characteristics are visualized. In this process the actual genetic composition (A,C,G, or T) is being measured. The composition/variant  in an offspring must correspond to the composition/variant in the mother and father that were provided for comparison. In two examples, it is shown how the basic rules are applied in parentage verification.

 

Marker

Offspring

Mother

Father

SNP01

AT

AA

TT

SNP02

GC

GC

CC

SNP03

TT

CT

TT

SNP04

AC

AC

AC

SNP05

CC

CC

CT

SNP06

CT

CC

CT

In the above table an example is provided of a correct parentage. In this table, the DNA is shown of three individuals: an offspring (left column), a potential mother (middle column), and a potential father (right columns). In each line one variant is shown. In this case all variants in the offspring are present in the parents: the parentage is correct.

 

Marker

Offspring

Mother

Father

SNP01

AA

AA

TT

SNP02

GC

GC

CC

SNP03

CC

CT

TT

SNP04

AC

AC

AC

SNP05

CC

CC

CT

SNP06

CT

CC

CT

In the second table an example is provided of an incorrect parentage. In this table, the DNA is shown of three individuals: an offspring (left column), a potential mother (middle column), and a potential father (right columns). In each line one variant is shown. In this case several variants are present in the offspring which are not are present in the parents: the parentage is not correct.

When 200 up to 400 different genetic fragments are checked, the chance that an incorrect parentage is not detected becomes very small. The genetic fragments which are used for parentage verification and identification provide no information on properties such as color and quality of an animal, plant or human, since the fragments are non-coding.

Useful information about DNA-profiles

The DNA-profile of one individual is identical in each part of the body. It does not make a difference in a comparison if a DNA-profile of an individual is based on hairs, blood, swabs, semen or tissue.

 Because large variation is present in the DNA, it is almost not possible that two randomly selected individuals have identical DNA-profiles. Each individual will have his or her own DNA, which will differ on one or more points from other individuals. One exception on this are identical twins or clones, which have completely identical DNA patterns.