Barbara Lillien
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South Shore H.S.
Summer
2001
WHAT
DOES THAT DNA MOLECULE REALLY LOOK LIKE???
SKILLS:
Extracting DNA from an animal cell
OBJECTIVES:
Separate and collect the DNA from liver cells
Describe the appearance of DNA extracted from a cell
Relate the location of DNA in a cell to procedures for extracting it
MATERIALS:
safety goggles
cheesecloth (several pieces, 12cm X 12 cm)
funnel
lab apron
beef liver ( 2 cm square/group of 2 students)
test
tubes
mortar & pestle
ice water bath
fine sand
test tube rack
graduated cylinder
70% Ethanol (4ml)
SDS/NaCl
solution(10ml)
inoculating loop
EDUCATION
AL
Unifying Concept:
Evidence, models, and explanation
Science &
Technology:
Understanding how science and technology come together
Life, Physical,
Earth Sciences: Molecular
basis of heredity, Structure and properties of matter
PURPOSE:
You are an intern
working in the city’s forensics lab. You
will be assisting the forensics technician with many of her routine laboratory
tests and procedures. One procedure
the technician does frequently is extract DNA from cells and purify it.
The purified DNA is used to help prepare a DNA fingerprint to help solve
crimes. To make sure you know how to do this procedure correctly, the
technician has asked you to extract DNA from the cells of a piece of beef liver
and spool the DNA for observation.
BACK
The extraction of
DNA from cells and its purification are of primary importance to the field of
biotechnology. Extraction and
purification of DNA are the first steps in the analysis and manipulation of DNA
that allow scientists to detect genetic disorders, p[produce DNA fingerprints of
individuals, and even create genetically engineered organisms used to produce
beneficial products such as insulin, antibiotics, and hormones.
The process of
extracting DNA, regardless of its original source, involves the following steps.
The first step in extracting DNA from a cell is to LYSE, or break open,
the cell. One common way to lyse cells is to grind a piece of
tissue along with a mild abrasive material in a mortar with a pestle.
After the cells have been broken open, a solution containing salt (NaCl)
and a detergent containing the compound SDS, or sodiumdodecyl sulfate, is used
to break down and emulsify the fat and proteins that make up the cell membrane.
Finally, ethanol is added. Because
DNA is soluble in water, the addition of ethanol causes the DNA to PRECIPITATE,
or settle out of solution, leaving behind all remaining cellular components that
are not soluble in ethanol. Finally,
the DNA can be wound (spooled) onto an inoculating loop, and pulled from the
test tube.
PROCED
1. Put on safety
goggles and lab coat
2. Place a piece of
liver in a mortar. Add several grains of sand.
3. Pour 10ml SDS/NaCl
solution in the mortar.
4. Use a pestle to
grind the ingredients until they form a thick fluid. (do not overgrind mixture)
5. Place several
layers of cheesecloth into a funnel. Pour the contents of the mortar through the
cheesecloth into a test tube until it contains at least 2 ml of the extract. (
gently squeeze the cheesecloth to remove all the fluid from the cheesecloth.)
6. Place the test
tube in an ice-water bath
7. Measure 4 ml of
ice-cold ethanol in a clean graduated cylinder
8. Hold the test
tube at a 45 degree angle.
a) slowly pour the
ethanol slowly down the side of the tube (do not pour the ethanol too fast or
directly into the liver solution)
b) as you pour the
ethanol into the test tube, observe the space between the ethanol and the liver
solution( the interface line), this is the DNA that you will be spooling onto
the inoculating loop.
9. Gently insert an
inoculating loop into the test tube as far as the interface line.
10 slowly move the
loop in circles, (this motion spools the long threads of DNA around the end of
the loop.
11. Lift the
inoculating loop out of the solution in the test tube.
While the DNA is being pulled out of the test tube, try stretching it.
Then dip the inoculating loop again to get more DNA.
12. When spooling is
complete, pull the inoculating loop from the test tube, and return the test tube
to a test-tube rack.
13. Clean up your
work area and wash your hands before leaving the lab.
QUESTIONS:
1. Describe the
appearance of the DNA you spooled from the crushed liver cells.
2. What was the
purpose of adding sand to the liver in the mortar?
3. What happens to
the cell when the SDS/NaCl solution is added to the liver mixture in the mortar?
4. What was the
purpose of filtering the liver mixture through cheesecloth?
5. What was done to
the DNA so that it could be observed and spooled?
6. How can you
determine whether the material pulled from the test tube was DNA?
7. How is DNA
protected inside an animal cell? How
does this location relate to the procedure you used in this lab to extract DNA?
8. Biotechnologists
research DNA. How could the procedure you used to day facilitate their research?
EXTENSIONS:
1. Find out what a
DNA fingerprint is and how it is used to compare samples of DNA from different
sources. Explain how the technique
you learned in today’s lab is used in developing a DNA fingerprint.
2. find out about the Human Gnome Project. What is this project attempting to do? How would the procedure used in this lab be used as part of the Human Genome Project