Deoxyribonucleic acid, most commonly known as DNA is one of the fundamental

biological molecule of a human beings and all other organisms whicg function mainly
but not limited to long-term storage of genetic information.
the objective of the experiment are to isolate the DNA from a plant source (Allium
cepa) and to determine the purity of the isolated DNA. the isolation of the DNA
from the plant source (Allium cepa) was made through disrupting its cellular
membrane and its other cellular components. based from the experiment, the isolated
DNA was found to have a thread-like structure. as for the test for purity of the
isolated DNA sample, the scientific technique that is spectroscopy was applied.
from the experiment, the DNA isolate had an absorbance of 3.631/3.251 which
indicates that the DNA isolate was contaminated.

spectropotometer reading values

grp7
a
230-3.853
260-3.634
280-3.261

b
230-3.844
260-3.628
280-3.240

ave 260-3.631 0.275

ave 280-3.2505 0.308

ratio-
3.631(260nm)/
3.251(280nm)
=1.12

Introduction

Nucleic acid is one of the fundamental biological molecule of an organism. nucleic

acids function as producer and storage of genetic information which directs the
process of protein synthesis, thus they are the one who determine which
characteristics a living thing would inherit. there are two principal type of
nucleic acids, these being deoxyribonucleic acid (DNA) and ribonucleic acid (RNA).
the experiment was mainly focused on DNA.

the structure of a nucleic acid include a nitrogenous base namely, purine or

pyrimidine. the base is covalently bonded to a 5- carbon sugar (ribose) on the 1',
while the sugar is covalently bonded to phosphate groups on their 5'.
as was previously mentioned, the nitrogenous bases of nucleic acids can be either
purine or pyrimidine. purines include adenine and guanine. these bond with sugars
on N9 covalently. on the other hand, pyrimidines are cystosine, uracil and thymine
wich covalently bons with the sugar on N7.

Deoxyribonucleic acid (DNA) functions mainly but is not limeted to long-term

storage of genetic information of an organism.other function of DNA includes
reproduction of genetical information of an organism through the process called
replication, which will be important for the diretcion and initiation of pro0tein
synthesis.

there is number of dissimilarity between DNA and RNA. First, DNA is known to have a
double helix structure whic means it has two strands that is oriented into a
antiparallel direction while RNA is made up of only one strand. Second, RNA's sugar
contains an OH bonded to its 2' carbon while DNA's sugar has only H. Lastly, DNA
and RNA differs in their adenine complementary base pairing, which in DNA, adenine
bonds with thymine. unlike DNA, RNA's adenine bonds with uracil.

the objective of the experiment are to (1) isolate DNA from a plant source (Allium
cepa) and (2) perform purity test using the scientific technique of ultraviolet
spectroscopy.

A. Compounds/ Sample used

Onion (Allium cepa), homogenizing solution: 5% SDS (sodium dodecyl sulfate or

sodium lauryl sulfate), 0.15 M NaCl, 0.15 M Na3C6H5O7 (sodium citrate), and 0.001 M
EDTA, Commercial papain or meat tenderizer (6% in water), ice cold 95% ethanol and
TE buffer

B. Procedure
1.Isolation of DNA from Allium cepa

50 ml of homogenizing solution was placed in a 125 ml erlenmeyer flask and was

subjected to water bath until it reached 60 degree celcius. 25g of minced Allium
cepa (onion) was added on the pre-heated homogenizing solution and was stirred and
let stood in a water bath form 5 minutes. 1.5g of papain was added and was kept
again in a water bath with 60 degree celcius for another 10 minutes. after this,
the flask was placed in an ice bath immediately and was swirled to prevent the
breakdown of the DNA. following the ice bath, the contents of the flask was placed
in a blender and was homogenized for 45 seconds. the product was filtered using a
four layered cheesecloth into a 250 ml beaker and. 15 ml of ice cold ethanol was
added by dripping it down on the side of the beaker producing a clear layer on top
of the filtrate. then, the DNA was precipitated because all the contents are
soluble to ethanol while DNA is not. the solution was let stood for 3-5 minutes.
bubbles was formed and the DNA was spooled out of the solution and was placed in to
a clean test tube.

2. Ultraviolet Spectroscopy

a 0.5 ml aliquot of DNA solution was dissolved in 4.5 ml of TE buffer. The solution
was transferred to a quartz cuvette and the absorbance was determined at 260 nm and
at 280 nm. the buffer solution was used as the blank solution. finally, the
A260/A280 ratio was calculated.

Result and Discussion

Isolation of DNA

the deoxyribonucleic acid (DNA) from the plant source, which was onion (Allium
cepa) went through three steps for it to be isolated, these steps include
homogenization, deproteinization and precipitation.

on the first part of the isolation which was the homogenization, the onion was
minced and was introduced to heat. the onion was treated with heat in order for its
cell wall to soften and for further cell breakdown. aditionally, heating results to
softening phospholipids present in the cell membrane of the onion. moreover, heat
denatures the enzyme, ribonuclease, which if existent will cause the fragmentation
of the DNA thereby making it difficult for the DNA from being spooled out.

after this was the deproteinization part wherein the protease enzyme named papain
was introduced to the solution. papain is a common enzyme which act as something
that would remove or prevent the protein which are present in the solution from
binding to the DNA. Lastly, addition of ice-cold ethanol was done for the
precipitation of the DNA. this process was done to make a separation of the DNA
component from the solution

its is also important to know, that the homogenizing solution played an important
role in the experiment. the solution is composed of three chemical namely, a
detergent which is called (1) Sodium Dodecyl Sulfate which acts in facilitating in
cellular membrane breakdown, (2) Ethylenediamine tetra acetic acid (EDTA) which
functions by weakening the cell through destablizing cations present in the cell.
(3) Sodium Chloride which assist in the separation and precipitation of the DNA
from the alcohol solution.

B. Ultaviolet Measurement of Isolated DNA

after the isolation of the DNA, the isolate was resuspended to a TE buffer to
undergo its test for purity through a process called ultraviolet spectroscopy. this
test will give an absorbance value which will be used as a comparison to a standard
apsortion value of a DNA to test its purity. in the results it was found that the
sample's absorbance was 000/00. this indicates that the sample is contaminated.

the calculation of the absobances value and ration allows quantitative measurement
of the DNA concentration and provides information about the degree of contamination
if present. DNA absorbs light most strongly at 260 nm, this is used as a comparison
to estimate the DNA concentration while the absorbance at 280 nm is used as an
indicator for protein concentration. A good quality that of whic is a pure DNA
sample should have an A260/ A280 ratio ranging from 1.7-2.0