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Şöhret Pektunç

CRIME SCENE INVESTIGATOR PCR BASICS

AIM: To find the real culprit by comparing the DNA samples taken from the crime
scene with the DNA samples of the suspects.

INTRODUCTION:
FORENSIC SCIENCE:
Word of forensic science was formed from the composition of two different Latin words,
Forensic Medicine and science. First of all, the forensic medicine is about a public
investigation and discussion. Examples of this are cases that were previously open to
the public. The second is that this word, which is reproduced from Greek for information,
is closely related to the scientific process which is the systematic method of obtaining
information today. The forensic science is used as a scientific process in crime solving
procedures. The forensic sciences originate from nearly every branch of science and
many views of modern society and are therefore considered to be an important part of
the forensic system.
Why is it necessary?
The forensic sciences; it provides important objective evidence as a result of the
evidence obtained in mortal cases, accidents or in any case. The forensic sciences are
also more important than the opinion of the eyewitness. (1)

Figure (1) is about the forensic science and it is adopted from:

https://leverageedu.com/blog/forensic-science-careers/
Human genetic structures have about 3 billion bases and 99.5% do not change. that is,
0.5% of the DNA sequence of humans is different and it is polymorphic sequences used
in forensic implementations.
As a result of the universal arragement, the DNA sequences used for judicial typing are
anonymous and are reproduced from parts of the chromosomes that do not check any
part so that they have no common properties or functions.(2)
SHORT TANDEM REPEAT:
A Short Tandem Repeat (STR) is a method of comparing specific loci on DNA with two
or more samples. The STR is a micro-satellite forming of two or thirteen nucleotides that
are repeated several or ten times in a row in the DNA helix. The STR detection plumbs
the number of sequences that repeat on DNA. The major difference between this
method and the polymorphism method is that it does not cut DNA with restriction
enzymes. This method is used to find the length of short tandem repeats depend on the
length of product produced by PCR.

Figure (2) is explaining the short tandem repeats and it is adopted from:
https://slideplayer.com/slide/8526371/
Uses of The Forensic;
The STR detection is an instrument that interprets specific STR regions where is in the
DNA. The versatile nature of the STR regions concentrates the difference between one
DNA profile and the other. The FBI-confirmed scientific instrument, STRmix, includes
this investigative method. The DNA profile system currently in use is depend on PCR
and it uses basic series or brief tandem repetition. This process is used in elevated
polymorphic zones which are short repetitive DNA series. STR loci are targeted with
series-special primers and enlarged using PCR. The STR loci are targeted with serial
specific primers and amplified using the PCR, the DNA pieces obtained are discreted
and stated using electrophoresis. (3)

Figure (3) is STR analysis and it is adopted from;

https://www.researchgate.net/figure/Principles-of-STR-analysis-STRs-loci-
comprise-repetitive-sequences-of-2-7-bp-which-are_fig1_26513043
MATERIALS:
1 Ice bath including tubes of DNA, 1 master mix+primers, 5 crime scene and suspect A-
D DNAs, 5 PCR tubes, 5 PCR adaptors,1 marking pen, 4 20 microliter adjustable
micropipette, 3 20 microliter tips, 3% agarose gel, 5 PCR sample from previous lab, 1
(300-350 ml) TAE running buffer, 60 microliter orange G loading dye, 25 microliter crime
scene investigator, gel electrophoresis chamber,power supply,fast blast DNA stain and
gel staining tray were used in this experiment.

METHODS:
20 microliter master mix + primers which were taken from the yellow tube with the help
of micropipette were placed into 5 PCR tubes. CS, A, B, C and D letters were written on
each different tubes for shouldn't mixed it. 20 microliter crime scenes were taken from
the purple tube and transferred to the CS tube. 20 microliter suspect A DNA were taken
from the greentube and transferred to the A tube. 20 microliter suspect B DNA were
taken from the blue tube and transferred to the B tube. 20 microliter suspect C DNA
were taken from the orange tube and transferred to the C tube. 20 microliter suspect D
DNA were taken from the pink tube and transferred to the D tube. Different micropipette
tips were used for each sample. Each sample was shaken with the mouth closed to
 ensure that it was mixed after being placed in the tubes. The PCR tubes are placed on
the ice adapters and then placed in the thermal cycler.
For Preparing %3 Agarose Gel;
3 g of agarose and 100 ml of TAE buffer were added to the flask to prepare the agarose
gel. This mixture, added to the flask, is placed in the microwave and is removed at
regular intervals, mixed and put back also this process was continued until the particles
are completely destroyed. When the particles disappeared completely, 2-3 drops of red
safe were added to the agarose gel mixture taken from the microwave and mixed. The
agarose gel was poured onto the tray for drying and the comb agarose gel was then
combed to form the vellers.
10 microliters orange g loading dye was placed in 5 PCR tubes and mixed well. The
dried agarose gel is placed on the gel electrophoresis apparatus and added the TAE
buffer until covered the agarose gel. 20 microliter Allele ladder is added with
micropipette starting from the 4 th box of the present agarose gel. 20 microliter Crime
Scene (CS), 20 microliter Suspect A, 20 microliter Suspect B, 20 microliter Suspect C,
and 20 microliter Suspect D were added with micropipette, respectively. The ends of the
PCR machine are connected to electricity and operated by setting at 100 V for 30
minutes.

RESULTS:
Allele Crime Suspect Suspect Suspect Suspect
Ladder Scene(CS) A B C D

10

1
Figure (4) is result of the experiment:
The bands formed as a result of electrophoresis are samples of persons who may be
guilty from the scene. The allele ladder in these results is used as reference for the
experiment. These bands were compared with the DNA of the other suspects. As a
result, the DNA samples taken from the crime scene match the bands manufactured by
the suspect C to a large extent. In this case, the suspect C is very likely to be guilty. The
electrophoresis and the bands can visualized under UV rays so can reach the suspect.
There are 3 main steps of this experiment. These are the PCR, electrophoresis and
visualizing the DNA under the ultra violet light. 20 microliter master mix + primers were
placed in each tube. 20 microliter of the samples taken from the criminals and suspects
were added to each different tube and mixed. Then 3 grams of agarose was added to
100 ml of TAE buffer and 3% agarose gel was prepared and allowed to dry and placed
in the PCR. The TAE buffer was poured until the agarose placed on the PCR was
covered. In the same time, orange G loading dye is added into the tubes to see the
DNA. In agarose gel, 20 microliters of each sample is placed in the wells, with space
between the wells. Then, the PCR machine is operated for 30 minutes at 100 voltages.
Finally, the agarose gel was examined under ultraviolet rays and the culprits were
detected by comparing the bands.

DISCUSSION:
The bands formed on the agarose gel as a result of the PCR were observed under
ultraviolet rays. As a result of the comparison of the bands according to the results, the
suspect C and the crime scene bands are the same, the suspect C and crime scene
DNA match and the culprit is the suspect C.
The result of the experiment is successful. Because one of the bands formed by the
samples matches that of the offender.
The above-described experiment did not produce errors; but may disintegration of the
agarose gel, secondly when placing the samples on the agarose gel with a micropipette,
if we push the micropipette button quickly and pull our hand, the sample in the box
passes through the other boxes and forms a band in double places, and lastly
contamination with any sample cause the errors.
The results of these experiments are used in forensic sciences to detect criminals or the
DNA of any individual can be compared.
Through STR loci, biological materials can be forensically identified, used in paternity
research, and specifically compare genetic markers.
As a result of performing the same formats as the DNA sample taken from the father
and child, paternity test can be performed and the experiment can be developed.
In conclusion of this experiment, the DNAs of the individuals were formed in bands and
the real culprit was easily identified as a result of comparison with the culprit's DNA.

REFERENCES:
1. International Student. (2019). What is Forensic Science? | Study Criminal and
Forensic Science in the US. [online] Available at:
https://www.internationalstudent.com/study-criminal-and-forensic-science/what-is-
forensic-science/ [Accessed 27 Oct. 2019].

2. Anon, (2019). [online] Available at: https://www.bio-rad.com/en-uk/product/crime-

scene-investigator-pcr-basics-kit?ID=58d47ce8-b4da-4c0c-9017-f217f27f27a4
[Accessed 28 Oct. 2019].

3. En.wikipedia.org. (2019). STR analysis. [online] Available at:

https://en.wikipedia.org/wiki/STR_analysis [Accessed 29 Oct. 2019].
FOR FIGURE;
1) Edu, L. (2019). An Overview of Forensic Science Careers And Programs-
Leverage Edu. [online] Leverage Edu. Available at:
https://leverageedu.com/blog/forensic-science-careers/ [Accessed 28 Oct. 2019].

2) Slideplayer.com. (2019). Commonly Used Short Tandem Repeat Markers - ppt

video online download. [online] Available at:
https://slideplayer.com/slide/8526371/ [Accessed 30 Oct. 2019].

3) Anon, (2019). [online] Available at:

https://www.researchgate.net/figure/Principles-of-STR-analysis-STRs-loci-
comprise-repetitive-sequences-of-2-7-bp-which-are_fig1_26513043 [Accessed 28
Oct. 2019].