Objective

To have a closer look and examine at the DNA molecule and extract our own
DNA using buccal cell.

Results

DNA

Discussion

Every living organisms, either plants or animal and even human, contain DNA.
Deoxyribonucleic acid (DNA) is the hereditary material in humans and almost all other
organismsis. DNA is a type of nucleic acid and it is made up of carbon, hydrogen, oxygen,

nitrogen, and phosphorous. It is the genetic material that an organism inherits from its
parents. Nearly every cell in a persons body has the same DNA. Most DNA is located in the cell
nucleus (where it is called nuclear DNA), but a small amount of DNA can also be found in the
mitochondria (where it is called mitochondrial DNA or mtDNA).Nucleotide is the building

blocks of nucleic acid. There are three main components which made up a nucleotide of
DNA, which are deoxyribose sugar, nitrogenous bases and phosphate group which carries
negative charge. There are four types of nitrogenous bases, which are adenine (A),
thymine (T), guanine (G) and cytosine (C). Deoxyribose sugar and phosphate group are
linked together by phosphodiester bond and they form the sugar-phosphate backbone of a
DNA molecule. DNA is formed by two polynucleotide strands which spiral around an
imaginary axis and forming a double helix. The two strands are complementary and
antiparallel with each other. Adenine always base pair with thymine, while guanine
always base pair with cytosine. There are two hydrogen bonds between adenine and
thymine. Guanine and cytosine are held together by three hydrogen bonds.
In this experiment, we are brought closer to look at our own human DNA. We extract our
own DNA from buccal cells as starting material. The whole experiment is carried out
with care to avoid any contamination. During the experiment, drinking water is used to
rinse a persons mouth to remove any food particles. Then, the person uses saline
mouthwash to swish his or her mouth and scrape the buccal cells off. If the person
swishes his or her mouth longer, more buccal cells can be removed and used to extract
DNA. After that, 10% (w/v) sodium dodecylsulphate (SDS) solution is added into the
centrifuge tube containing saline solution and buccal cells. Thermo Scientific Sodium
Dodecyl Sulphate (Lauryl) is standard-grade SDS detergent. It is normally used for
denaturing protein polyacrylamide gel electrophoresis (PAGE). SDS is a strong anionic
detergent which is used for routine protein electrophoresis and cell lysis methods.
Besides, it is also a common component of cell lysis buffers. Detergents are commonly
used to lyse cells, solubilize membrane proteins and lipids, control protein crystallization,
prevent nonspecific binding in affinity purification and immunoassay procedures, and as

additives in electrophoresis. When SDS detergent is added, it removes negative ions from
the protein of the cell membranes of buccal cells causing the protein to damage. SDS
disrupts the cell membranes and nuclear envelopes of buccal cells and destabilise the
hydrophobic interactions among the macromolecules. The buccal cells burst opem and
DNA is released. After that, Bromelain protease enzyme is added into the centrifuge tube.
Bromelain is a mixture of proteolytic enzymes and other substances in small quantities.
Proteolytic enzymes are sulfhydryl proteases as a free sulfhydryl group of a cysteine sidechain is required for function. It catalyses the breakdown of protein into amino acids
through hydrolysis process. DNA is wrapped very tightly around proteins which are
histones in the nucleus, so bromelain protease enzyme is used to remove these proteins
from DNA to free DNA. After that, the centrifuge tube is put into a warm water bath for
10 minutes. The time must be long enough for DNA to free from the cells. Then, ice cold
isopropanol is slowly added into the centrifuge tube to precipitate out DNA from the
solution. This is because DNA is less soluble in isopropanol and it will come out of the
solution. Besides, isopropanol also allows precipitation of larger species and lower
concentrations of nucleic acids than ethanol. Isopropanol added must be cold because
DNA is more stable in cold temperature. DNA is heat- sensitive and it is denatured easily
at high temperature. Salts in the solution are less soluble in isopropanol, so they have
more tendencies to co-precipitate with the DNA. Isopropanol precipitations are carried
out at room temperature with minimal incubation times to lessen the chances of salt
precipitation.
DNA that we extract appeared as a colourless jelly-like substance. It looks whitish. In a
very real sense, each of us know that the shape of DNA is a long double helix structure,
but the one we extract from this experiment is in smaller fragments. In short, the
appearances of our DNA are colourless, small, coiled together and float at the upper part
of the solution.The extracted DNA can be seen with naked eyes and easily. It looks like
strands of mucous-like, translucent cotton. The extracted DNA consists of thousands of
DNA molecules which stuck together with some proteins too. When DNA is released
from the buccal cells, it breaks up into tiny fragments by nuclease enzyme. These tiny
DNA fragments carry slightly negative electric charge. Salt ions in the solution are

attracted to the negative charges on the DNA fragments and prevent them from adhering
to one another. So, DNA molecules will not clump together.
The extracted DNA can be used for genetic fingerprinting. Everyone has their own
different and unique DNA, although close relatives can have some similarities in their
DNA. Monozygotic twins cannot be identified by genetic fingerprinting as their DNA
sequences are totally the same. Genetic fingerprinting is a process in which an
individual's unique sequence of DNA base pairs can be determined by exposing a sample
of the person's DNA to molecular probes. DNA fingerprints are always used as evidence
in criminal law cases. Forensic scientists use DNA fingerprinting to identify a criminal or
victim in a criminal investigation based on their unique DNA profiles. DNA profiles are
encrypted sets of numbers that reflect a person's DNA makeup and acts as his or her
identifier. DNA profiling is different from full genome sequencing. DNA fingerprinting is
commonly used in forensics to exonerate innocent people whove been wrongly
convicted in a crime, resolve paternity disputes, to determine genetic characteristics of
offspring including the diagnosis of diseases in the offspring and to identify an
abandoned offspring, identify the victims in cases where they cannot be identified by
using the dental records, to determine the pedigree for livestock or pets, and identify
carriers of defective genes. In forensics, DNA can be obtained from blood, hair follicles,
body tissues, fluids and others. Then, the DNA obtained can be matched with a sample
obtained from a pool of suspects in DNA fingerprinting to identify the criminal.
Sometimes, DNA is used in wildlife forensics to identify endangered species and
poachers. DNA is also used in cloning process to produce multiple clones of specific
animal or plant species.
There are several precautions in this experiment. First, do not touch the tip of the pipette
to the inside of the centrifuge tubes or the samples may become contaminated. Vortex is
used to mix the solution thoroughly. Furthermore, test tube containing Isopropanol must
be immersed in the ice throughout the experiment to ensure that it is always cold. The tip
of the pipette filling with ice cold isopropanol must put against the inside wall of the
centrifuge tube when isopropanol is added into the tube. Add isopropanol slowly to
precipitate out DNA from the solution successfully. Besides, after isopropanol is added

into the centrifuge tube, does not mix, shake, or bump the test tube when it is placed in
the test tube rack for 10 minutes. This is to make sure that DNA does not shear and can be
extracted successfully. If DNA is broken or sheared in too many places, it wont spool
and is harder to capture effectively. DNA should be precipitated out in white or clear
clumps that may also look like cobwebs or gooey strands.
Conclusion
DNA can be extracted from the human buccal cells. The extracted DNA looks like
strands of mucous-like, translucent cotton and can be seen easily with naked
eyes. The extracted DNA can be used in various fields for benefits of the
community and country.
Question
1. Which one of the following do you think will contain DNA?
Bananas; concrete; fossils; meat; metal; spinach; strawberries.
Explain your reasoning.
Bananas, fossils, meat, spinach, and strawberries. Because all of them are living
things, while the others are non-living things. Cells are the basic building blocks
of all living things and cell contains of nucleus and other organelles. The nucleus
serves as the cells command centre, sending directions to the cell to grow,
mature, divide, or die. It also houses DNA (deoxyribonucleic acid), the cells
hereditary material. The nucleus is surrounded by a membrane called the nuclear
envelope, which protects the DNA and separates the nucleus from the rest of the
cell.
2. What effect would the SDS have on the cell membranes and cold ethanol on
DNA?
SDS is a detergent which causes the cell membrane to break down further and
emulsifies the lipids and proteins of the cell by disrupting the polar interactions
that hold the cell membrane together. The detergent forms complexes with these
lipids and proteins causing them to precipitate out of the solution. Using ice-cold

water and ice-cold alcohol will increase your yield of DNA. The cold water
protects the DNA by slowing down enzymes that can break it apart. The cold
alcohol helps the DNA precipitate (solidify and appear) more quickly.
3. What type of enzyme would be needed to separate the DNA into smaller pieces?
Restriction enzymes
4. Is the DNA extracted pure enough for further applications (i.e. PCR)?
No, because the DNA obtained from buccal cell might be contaminated.
5. If you were to repeat the experiment with an equal number of red blood cells, the
amount of DNA collected would either: increase / decrease / stay the same
(choose one). Explain your answer.
Decrease. Although blood is an excellent source of DNA, the DNA does not come
from the red blood cells, as these cells have no nuclei. Rather, the DNA comes
chiefly from white blood cells in the blood.