Isolation of RNA from Yeast

Dizon, Allen Sergio D.R. 2FMT

Faculty of Pharmacy,
Department of Medical Technology,
University of Santo Tomas

ABSTRACT
Ribonucleic acid was isolated from yeast by heating the active dry yeast with 1% NaOH
solution. The RNA was then obtained by separating it from proteins by adding HCl by utilizing
acid extraction together with ethanol to precipitate the RNA molecules out of the aqueous
solution. To determine the purity of the isolated RNA, its absorbance was measured at 260nm
and at 280nm that yielded a result of 0.218 which confirmed that the RNA sample that was
isolated is impure and that it contains possible contaminants.

The main objectives of the study is to [1]

INTRODUCTION
Ribonucleic acid (RNA) molecules are single-
stranded polymer nucleic acid made up of
ribonucleotides. Each ribonucleotide consists of
a ribose sugar, a phosphate, and a nitrogenous
base derived from purines which are adenine
(A) and guanine (G) and from pyrimidines which
are cytosine (C) and uracil (U). In comparison
with DNA, RNA is relatively shorter in molecules
thus it is not easily damaged by shearing but it
is more unstable and more prone to
degradation due to having a ribose sugar.
Figure 1. Structure of a Nucleotide
The RNA is
Isolate RNA from the yeast sample [2]
Calculate the percent yield of the obtained
RNA. [3] Measure the isolated RNA through
Ultra Violet Measurement.
METHODOLOGY
A. Isolation of RNA
3.0g of active dry yeast was dissolved into
25mL of water with 5mL of 1% NaOH
solution. The dry yeast solution was heated to
60c water bath for 15 minutes while stirring
occasionally. After heating, the solution was
strained through a cheesecloth. The filtered
solution was centrifuged and then glacial
acetic acid was added to the supernate until
faintly acidic to the litmus paper. The
suppernate was evaporated over a water bath
until 10mL of liquid remains. The supernate
was cooled to 40c and was then poured to a
solution of 20mL of 95% alcohol and 0.2mL
of concentrated HCL. The solution was
stored afterwards to let the RNA settle at the
bottom of the test tube. After the RNA have
settled, it was decanted and washed twice
with 5mL of 95% ethanol and twice again
with ether.
B. Ultraviolet Measurement of RNA
The aliquot of RNA was diluted with TE
buffer and was mixed. The solution was
transferred to was transferred to a microwell
plate and the absorbance was measured at
260nm and at 280m.
RESULTS AND DISCUSSION
theoretically a blue solution must be seen as
a positive result in the deoxyribose test, a red
residue in the test for purines, and a purple
coloration in the test for pyrimidines. Also the
errors in the DNA characterization was also
due to the improper execution of the
experimentalists.

A. Isolation of RNA and UV Measuremnet

REFERENCES
Ribonucleic Acid. (n.d.). In Scitable online.
Table 1. RNA Isolation results Retrieved from
http://www.nature.com/scitable/definition
RNA Sample Physical Description /ribonucleic-acid-rna-45

Bowen, R. (2001, Dec 9). The Structure of

Yeast White Substance Nucleic Acids.Retrieved from
http://arbl.cvmbs.colostate.edu/hbooks/g
enetics/biotech/basics/nastruct.html
Table 2. Quantitative Analysis of RNA
UV Measurement [2] Ribonucleic Acid. (n.d.). In Scitable online.
Total RNA Retrieved from http://www.nature.com/scita
(A260/A280) (A260)/A230)

0.218 - 1,744mg

The isolated RNA had an appearance of

white substance that settles at the bottom of
the container. The calculated purity of RNA
is about 0.218 which means that the isolated
RNA sample is not pure and that
contaminants are

CONCLUSION

DNA hydrolysis was done to separate

the components of nucleic acid. DNA reacts
in acid hydrolysis and is less reactive in basic
hydrolysis since it is negatively charged
because of the phosphate surrounding it.

There are erroneous color results in

the standards in the chemical
characterization because of the reagent
used. The reagents might not have been
used in a long time. Because the test for
deoxyribose yielded a negative standard but
the DNA sample yielded the same negative
result which makes the result positive,