ISOLATION OF DNA FROM

VARIOUS FRUITS
ACKNOWLEDGEMENT
I feel proud to present my Investigatory
project in Biology on the topic
ISOLATION OF DNA FROM DIFFERENT
FRUITS which aims at seprating DNA
from various types of fruits.This project
wouldnt have been feasible without the
proper and rigorous guidance of my
biology teacher Mrs. Vimala Jossy who
guided me throughout this project in
every possible way.
INDEX
INTRODUCTION
CHROMOSOMES
A chromosome is a packaged and organized structure
containing most of the DNA of a living organism..
Most eukaryotic cells have a set of chromosomes (46
in humans) with the genetic material spread among
them.
During most of the duration of the cell cycle, a
chromosome consists of one long double-helix DNA
molecule (with associated proteins). During S phase,
the chromosome gets replicated, resulting in an Xshaped structure called a metaphase chromosome.

Both the original and the newly copied DNA are now
called chromatids. The two "sister" chromatids are
joined together at a protein junction called a
centromere (forming the X-shaped structure).
Chromosomes are normally visible under a light
microscope only when the cell is undergoing mitosis
(cell division). Even then, the full chromosome
containing both joined sister chromatids becomes
visible only during a sequence of mitosis known as
metaphase (when chromosomes align together,
attached to the mitotic spindle and prepare to
divide).This DNA and its associated proteins and
macromolecules is collectively known as chromatin,
which is further packaged along with its associated
molecules into a discrete structure called a
nucleosome. Chromatin is present in most cells, with
a few exceptions, for example, red blood cells.
Occurring only in the nucleus of eukaryotic cells,
chromatin composes the vast majority of all DNA,
except for a small amount inherited maternally, which
is found in mitochondria.
In prokaryotic cells, Chromatin occurs free-floating in
cytoplasm, as these cells lack organelles and a
defined nucleus. The main information-carrying
macromolecule is a single piece of coiled double-helix
DNA, containing many genes, regulatory elements
and other noncoding DNA. The DNA-bound
macromolecules are proteins that serve to package
the DNA and control its functions. Chromosomes vary
widely between different organisms. Some species
such as certain bacteria also contain plasmids or
other extrachromosomal DNA. These are circular
structures in the cytoplasm that contain cellular DNA

and play a role in horizontal gene transfer.

Compaction of the duplicated chromosomes during
cell division (mitosis or meiosis) results either in a
four-arm structure (pictured above) if the centromere
is located in the middle of the chromosome or a twoarm structure if the centromere is located near one of
the ends. Chromosomal recombination during meiosis
and subsequent sexual reproduction plays a
significant role in genetic diversity. If these structures
are manipulated incorrectly, through processes
known as chromosomal instability and translocation,
the cell may undergo mitotic catastrophe and die, or
it may unexpectedly evade apoptosis leading to the
progression of cancer.
In prokaryotes (see nucleoids) and viruses, the DNA is
often densely packed and organized: in the case of
archaea, by homologs to eukaryotic histones, and in
the case of bacteria, by histone-like proteins. Small
circular genomes called plasmids are often found in
bacteria and also in mitochondria and chloroplasts,
reflecting their bacterial origins.
Deoxyribonucleic acid is a molecule that carries the
genetic instructions used in the growth, development,
functioning and reproduction of all known living
organisms and many viruses. DNA and RNA are
nucleic acids; alongside proteins, lipids and complex
carbohydrates (polysaccharides), they are one of the
four major types of macromolecules that are essential
for all known forms of life. Most DNA molecules
consist of two biopolymer strands coiled around each
other to form a double helix.
The two DNA strands are termed polynucleotides

since they are composed of simpler monomer units

called nucleotides. Each nucleotide is composed of
one of four nitrogen-containing nucleobaseseither
cytosine (C), guanine (G), adenine (A), or thymine (T)
and a sugar called deoxyribose and a phosphate
group. The nucleotides are joined to one another in a
chain by covalent bonds between the sugar of one
nucleotide and the phosphate of the next, resulting in
an alternating sugar-phosphate backbone. The
nitrogenous bases of the two separate polynucleotide
strands are bound together (according to base pairing
rules (A with T, and C with G) with hydrogen bonds to
make double-stranded DNA. The total amount of
related DNA base pairs on Earth is estimated at 5.0 x
1037, and weighs 50 billion tonnes. In comparison,
the total mass of the biosphere has been estimated to
be as much as 4 trillion tons of carbon (TtC).
DNA stores biological information. The DNA backbone
is resistant to cleavage, and both strands of the
double-stranded structure store the same biological
information. This information is replicated as and
when the two strands separate. A large part of DNA
(more than 98% for humans) is non-coding, meaning
that these sections do not serve as patterns for
protein sequences.
The two strands of DNA run in opposite directions to
each other and are thus antiparallel. Attached to each
sugar is one of four types of nucleobases (informally,
bases). It is the sequence of these four nucleobases
along
the
backbone
that
encodes
biological
information. RNA strands are created using DNA
strands as a template in a process called
transcription. Under the genetic code, these RNA

strands are translated to specify the sequence of

amino acids within proteins in a process called
translation.

Within eukaryotic cells, DNA is organized into

long structures called chromosomes. During
cell division these chromosomes are duplicated
in the process of DNA replication, providing
each
cell
its
own
complete
set
of
chromosomes. Eukaryotic organisms (animals,
plants, fungi, and protists) store most of their
DNA inside the cell nucleus and some of their
DNA in organelles, such as mitochondria or
chloroplasts.[6]
In
contrast,
prokaryotes
(bacteria and archaea) store their DNA only in
the
cytoplasm.
Within
the
eukaryotic
chromosomes, chromatin proteins such as
histones compact and organize DNA. These
compact structures guide the interactions
between DNA and other proteins, helping
control which parts of the DNA are transcribed.
DNA was first isolated by Friedrich Miescher in 1869.
Its molecular structure was identified by James
Watson and Francis Crick in 1953, whose modelbuilding efforts were guided by X-ray diffraction data
acquired by Rosalind Franklin. DNA is used by
researchers as a molecular tool to explore physical
laws and theories, such as the ergodic theorem and
the theory of elasticity. The unique material
properties of DNA have made it an attractive
molecule for material scientists and engineers

interested in micro- and nano-fabrication. Among

notable advances in this field are DNA origami and
DNA-based hybrid materials.
METHODOLOGY
DNA isolation is a process of purification of DNA from
sample using a combination of physical and chemical
methods. The first isolation of DNA was done in 1869
by Friedrich Miescher.Currently it is a routine
procedure in molecular biology or forensic analyses.
BASIC PROCEDURE
There are three basic and two optional steps in a DNA
extraction:

Cells which are to be studied need to be

collected.

Breaking the cell membranes open to expose the

DNA along with the cytoplasm within (cell lysis).

Lipids from the cell membrane and the nucleus

are broken down with detergents and
surfactants.

Breaking proteins by adding a protease .

Breaking RNA by adding an RNase.

The solution is treated with concentrated salt

solution to make debris such as broken proteins,
lipids and RNA to clump together.

Centrifugation of the solution, which separates

the clumped cellular debris from the DNA.

DNA purification from detergents, proteins, salts

and reagents used during cell lysis step. The

most commonly used procedure is:

Ethanol precipitation usually by ice-cold ethanol

or isopropanol. Since DNA is insoluble in these
alcohols, it will aggregate together, giving a
pellet upon centrifugation. Precipitation of DNA
is improved by increasing of ionic strength,
usually by adding sodium acetate.

Cellular and histone proteins bound to the DNA can be

removed either by adding a protease or by having
precipitated the proteins with sodium or ammonium
acetate, or extracted them with a phenol-chloroform
mixture prior to the DNA-precipitation.
After isolation, the DNA is dissolved in slightly
alkaline buffer, usually in the TE buffer, or in ultrapure water.
TO ISOLATE DNA FROM VARIOUS TYPES OF FRUITS.
Rubbing alcohol, Measuring cup,Measuring
spoons,Salt,Water,Dishwashing liquid (for handwashing dishes),Glass or small
bowl,Cheesecloth,Funnel,Tall drinking glass,Three
fruits,Resealable plastic sandwich bag,Small glass jar
(such as a spice or baby food jar),Bamboo skewer,
available at most grocery stores. (If you use a baby
food or short spice jar, you could substitute a
toothpick for the skewer.)
PREPARATION

Chill the rubbing alcohol in the freezer.

Mix one half teaspoon of salt, one third cup of

water and one tablespoon of dishwashing liquid
in a glass or small bowl. Set the mixture aside.

This is your extraction liquid. Why do you think

there is detergent in the extraction liquid

Completely line the funnel with cheesecloth.

Insert the funnel tube into the tall drinking glass
(not the glass with the extraction liquid in it).

Remove and discard the green tops from the

fruits.

PROCEDURE

Put the fruits into a resealable plastic

sandwich bag and push out all of the extra air.
Seal the bag tightly.

With your fingers, squeeze and smash the

fruits for two minutes.

Add three tablespoons of the extraction liquid

you prepared to the fruits in the bag. Push out
all of the extra air and reseal the bag.

Squeeze the fruits mixture with your fingers

for one minute.

Pour the fruits mixture from the bag into the

funnel. Let it drip through the cheesecloth and
into the tall glass until there is very little liquid
left in the funnel (only wet pulp remains).

Pour the filtered fruits liquid from the tall glass

into the small glass jar so that the jar is one
quarter full.

Measure out one half cup of cold rubbing

alcohol.

Tilt the jar and very slowly pour the alcohol

down its side. Pour until the alcohol has

formed approximately a one-inch-deep layer on
top of the fruits liquid. You may not need all of
the one half cup of alcohol to form the one-inch
layer. Do not let the fruits liquid and alcohol
mix.

Study the mixture inside of the jar. The fruits

DNA will appear as gooey clear/white stringy
stuff.

CALCULATIONS - Extra: If you have access to a

milligram scale (called a balance), you can measure
how much DNA you get (called a yield). Just weigh
your clean bamboo skewer and then weigh the
skewer again after you have used it to fish out as
much DNA as you could from your fruit DNA
extraction. Subtract the initial weight of the skewer
from its weight with the DNA to get your final yield
of DNA.

OBSERVATIONS
While other fruits are soft and just as easy to pulverize,
strawberries are the perfect choice for a DNA extraction lab
for two very good reasons: (1) they yield way more DNA
than other fruits, and (2) they are octoploid, meaning that
they have eight copies of each type of DNA chromosome.
(Human cells are generally diploid, meaning two sets of
chromosomes.) These special circumstances make
strawberry DNA both easy to extract and to see.
When you added the salt and detergent mixture to the smashed
strawberries, the detergent helped lyse (pop open) the strawberry
cells, releasing the DNA into solution, whereas the salt helped
create an environment where the different DNA strands could
gather and clump, making it easier for you to see them. (When you
added the salt and detergent mixture, you probably mostly just saw
more bubbles form in the bag because of the detergent.) After you

added the cold rubbing alcohol to the filtered strawberry liquid, the
alcohol should have precipitated the DNA out of the liquid while the
rest of the liquid remained in solution. You should have seen the
white/clear gooey DNA strands in the alcohol layer as well as
between the two layers.
To extract the DNA, each component of the extraction
mixture plays a part. Soap helps to dissolve cell membranes.
Salt is added to release the DNA strands by breaking up
protein chains that hold nucleic acids together. Finally, DNA
is not soluble in isopropyl alcohol, especially when the
alcohol is ice cold.