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DNA from
Whole Blood
Group 4 - 4Bio2
Binag, Sam Dominic
Caringal, Joe Mari Isabella
Chiang, Rowena
Verin, Maryneil
Introduction
DNA EXTRACTION
process of purification of DNA from sample
using a combination of chemical and
physical methods
one of the most widely used molecular
biology techniques
usually the first step in a longer laboratory
process
Introduction
DNA EXTRACTION
important routine procedure in the fields of
biotechnology and forensics
allow scientists to detect genetic diseases,
analyze forensic evidence, produce DNA
fingerprint of individuals, and create of
genetically engineered organisms that can
produce beneficial products.
Introduction
DNA EXTRACTION OVERVIEW
1. lyse or break open the cell to expose the
DNA of interest.
Introduction
BLOOD
excellent source of human DNA
Introduction
Blood
Whole blood contains three types of
cells: red blood cells (RBCs), white
blood cells (WBCs), and platelets.
RBCs do not have any DNA
Objectives
To prepare a flow diagram of the
experimental protocol
To extract DNA from blood
Hypothesis
Methodology
Results and
Discussion
Cell Concentration
Centrifugation
Phosphate Buffered Saline (PBS)
Cell Lysis
STE, SDS, Proteinase K
Results and
Discussion
STE
Sodium chloride: for DNA to precipitate
Tris: buffer system
EDTA: chelates cations
SDS
Detergent
Disruption of cell membranes
Emulsification of lipids and proteins
Results and
Discussion
Proteinase K
Denatures proteins
Disables nucleases
DNA Extraction
Phenol-chloroform extraction
Liquid-liquid extraction
For isolation of DNA, RNA, proteins
Results and
Discussion
Forms lower organic phase and upper
aqueous phase
Results and
Discussion
Aqueous Phase
Contains hydrophilic DNA
Organic Phase
Contains hydrophobic lipids
White interphase
Interactions of hydrophobic molecules of proteins
with phenol
Results and
Discussion
Phenol: constitutes lower organic
phase
Chloroform
Miscible with phenol
Forces a sharper separation between
organic and aqueous phase
Isoamyl alcohol
Added in small amounts
Prevents foaming
Results and
Discussion
DNA Concentration (Washing)
Sodium acetate (Salt)
Makes DNA less soluble in the aqueous solution
More efficient precipitation of DNA in alcohol
Ethanol Precipitation
Washing with 70% Ethanol
Wash away residual salts
Results and
Discussion
TE Buffer
Tris-EDTA Buffer
Solubilize DNA while preventing
degradation
Guide Questions
1. What is the role of the following in DNA
extraction?
a. Ethanol
b. NaCl
c. SDS
d. TE Buffer
e. EDTA
Guide Questions
a. Ethanol
Ethanol is used in DNA extraction to
force the DNA to precipitate in a solution.
Ethanol prevents the DNA from dissolving
into the water, instead causing it to
precipitate out so it can be separated
and extracted. Without the addition of
ethanol, DNA will just dissolve in water.
Guide Questions
b. NaCl
Guide Questions
c. SDS
Guide Questions
d. TE Buffer
Guide Questions
e. EDTA
Guide Questions
2.Give the complete chemical names of the
following:
a. SDS Sodium dodecyl sulfate
b.Tris BufferTris(Hydroxymethyl)aminomethane
Guide Questions
3. Why should the extracted DNA be
immersed in TE buffer?
TE buffer protects DNA from
degradation.
"TE"
stands
for
its
components Tris and EDTA. Tris serves as
a buffering agent while EDTA serves as a
chelating agent. TE buffer is commonly
used to protect the DNA while storing it.
Conclusion
The purpose of DNA extraction is to obtain DNA
in a relatively purified form, which can be used
for further investigations.
An efficient DNA extraction method for whole
blood was carried out using phenol-chloroform
extraction process.
The protocol designed for blood yielded DNA.
The number, age, and freshness of the source
affect the quality and quantity of the DNA
obtained.
Thank you!