LAB REPORT #3: DNA + RNA

Biology I, Pre-Health Sciences – Durham College, Fall 2020

/40 marks
EVALUATION NOTES

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 While students may work in groups to complete labs, each student must submit
their own original work.
PART 1 – DNA EXTRACTION

Introduction
To most biology students,
deoxyribonucleic acid (DNA) feels a bit
abstract. It is common for many to know
that DNA has a ‘double helix’ and is the
‘genetic code’ for life, but beyond that it is a
bit hard to conceptualize.
The purpose of this part of the laboratory is
to demonstrate the process of isolating
DNA from various plant and animal tissues.
The process starts with a whole tissue and
ends with a relatively pure preparation of
DNA, containing literally billions of genes.
By observing DNA in this way, it will
become less of a ‘strange and mysterious
substance’ and more of a tangible, practical
molecule that holds the key to an
organism's development and structure.
Note: The procedure is described here so that you can replicate the experiment at
home if you choose to. However, you do not need to do so in order to complete the lab
report questions. A demo of the experiment is provided so you have everything you
need to answer the questions.

Materials Used:
 Cell samples – 2-3 strawberries (fresh or frozen); ½ of a banana can be used as
an alternative
 Equipment – small sieve or coffee filter or cheesecloth, 250 mL measuring cup,
butter knife or single chopstick, 2 glass cups, kitchen measuring spoons
(teaspoons, tablespoons), timer/clock, toothpicks
 Buffer ingredients – sodium chloride (salt; NaCl), sodium bicarbonate (baking
soda; NaHCO3), detergent (dish soap)
 Separation reagent – ice cold 70% isopropanol (rubbing alcohol)
Procedure

1. First, a sample of cells must be obtained. Cells can come from many places in
our everyday lives but for this lab we will focus on the extraction of DNA from
strawberry cells. There are 2 reasons for this: Strawberry cells are polyploid –
this means that they have lots of copies of their DNA in the nucleus. This makes it
easier to extract large quantities of DNA. The other reason is more practical –
strawberries are soft and easy to mash up.

a. To prep the strawberry cells: Add 2-3 strawberries to a glass cup. Using
the blunt end of a butter knife or chopstick, mash the strawberry until it is a
very creamy consistency (like a strawberry milk shake!). Some versions of
this experiment use a Ziplock bag to mash the strawberries in, but this one
is a more environmentally-friendly version.

Figure 3.1 Image of mashed strawberry. This image

contains more strawberry than needed for the experiment,
but gives an idea of how mashed they should be.

Fun Fact: In addition to strawberry cells, you could also collect a sample of cells
from the inside of your own cheek. To do this, you would need to prepare a 0.1%
NaCl solution (dissolve about 1 teaspoon of table salt in 1 liter of warm water) and
swirl one mouthful of (about 10 milliliters) for 2 minutes. Then, spit the salt solution
into one of the glass cups. DNA extraction from cheek cells is a bit more difficult as
there are fewer starting cells in comparison to the strawberry, but you should still
be able to extract a small quantity of your own DNA.

2. Next, make 50 mL (1/4 cup) of buffer solution. Measure all ingredients as

indicated below, and mix thoroughly in a 250 mL measuring cup until all
components are dissolved.

Hot tap water ¼ cup

Salt 1 teaspoon
Baking soda 1 teaspoon
Detergent (dish soap) ½ teaspoon
3. Next, combine the buffer and the cells. Add about 20 mL (1.5 tablespoons) of
buffer solution to the sample of strawberry cells and about 20 mL (1.5
tablespoons) of buffer solution to the sample of cheek cells (if they were collected).
Stir this mixture vigorously for 3 minutes using the butter knife/chopstick. NOTE:
During this time, each of the buffer ingredients play an important role in the DNA
extraction process:

 The detergent is used to break down the “greasy” membranes present

within the cell. By destroying the nuclear membrane and the plasma
membrane, the contents of the cell (including the DNA) can escape into the
solution.
 The salt (NaCl) and baking soda (NaHCO 3) are used to adjust the salinity
(salt concentration) and pH (acidity) of the solution, respectively. This helps
keep the DNA molecules stable so that they are less likely to break down.

4. Set up a simple filtration apparatus using a small sieve (or coffee filter twisted
into a cone shape or a square of cheesecloth) placed on top of a clean glass cup.
Filter the cell/buffer solution through the filter. Collect as much filtrate (liquid in the
cup) as possible. Dispose of any leftovers (solids in the sieve) in the compost
bin.

Figure 3.2 Filtration apparatus for isolation of strawberry cells. A

coffee filter or cheesecloth may be used instead.

5. Next, it’s time to separate the DNA from the filtrate. Measure out about 20 mL (1.5
tablespoons) of ice-cold 70% isopropanol (rubbing alcohol) using your kitchen
measuring spoons. Gently, add the isopropanol to your filtrate. The isopropanol
causes 2 layers (phases) to form in the glass. The bottom layer is aqueous and
contains the cellular components that are able to dissolve in water. The top layer
is non-aqueous and contains the cellular components that are not able to dissolve
in water.
 IMPORTANT: It is very important that you do not disturb the 2 layers. This
means NO stirring or shaking!

6. Wait 10-15 minutes. BE PATIENT. DNA will slowly begin to form where the
aqueous and non-aqueous layers meet, and will likely float to the top of the
isopropanol (non-aqueous) layer.

(1.5 marks) Based on the descriptions provided in steps 5 and 6, label the
aqueous layer, non-aqueous layer, and DNA on the following schematic
of the extraction:
 A. ____NON-AQUEOUS LAYER_______

B. __________DNA__________________

C. __AQUEOUS LAYER____________

7. Use toothpicks to spool the DNA for closer observation.

If you were in a laboratory and the DNA sample needed

to be collected and stored, it would be extremely
important to put a proper label on the collection
container. A proper label should include: your name,
the date, and the type of DNA collected (strawberry or
cheek).

(1.5 marks) Write out an example of a proper label for

the DNA sample if you had collected it in the lab today.

Tracey Bowles, October 9, 2020, Strawberry DNA

Figure 3.3 Strawberry DNA spooled with a
wooden skewer.

8. (2 marks) Would this experiment produce DNA if bananas were used instead of
strawberries? Explain why or why not. Yes, this experiment would produce DNA if
bananas were used instead of strawberries. All living things contain DNA.
Therefore, provided you can “mash” it up to examine the sample in this fashion,
this experiment would work.

9. (3 marks) A buffer solution (Step 2 and 3 above) was used to extract DNA from a
 strawberry. Using the table below, identify the role that each of the components
plays during the DNA extraction process, i.e. why was each component needed?
Remember to answer in your own words!

Buffer Component Role in DNA Extraction Process

Salt (NaCl) is used to adjust the salinity of the buffer solution, and is
Salt (NaCl) used in conjunction with the Baking Soda, to maintain the stability of
the DNA and prevent breakdown.
Baking Soda Baking Soda (NaHCO3) helps to stabilize the DNA by controlling the
(NaHCO3) PH levels of the buffer solution.
The dish soap is used to break down the phospholipid membranes
Dish soap within the cells.

10. (1 mark) If you were to extract DNA from your own cheek cells, how might the end
result differ from that of the strawberry DNA? The main difference to the end
result, of an experiment on cheek cells, would be that there would be much less
DNA to examine.
11. The extraction of nucleic acids plays a vital role in health-care and scientific
research settings.

To start, the diagnosis of many diseases can be done by coupling DNA extraction
with DNA sequencing to look for mutations in the genetic code. For example, a
genetic mutation in the BRCA1 gene can be identified using DNA extraction and
DNA sequencing so that women with breast cancer can be best treated. Prenatal
testing is another common use of DNA extraction. As fetal DNA is present in small
amounts within the mother’s blood, fetal DNA can be extracted from a maternal
blood sample and tests can be run to determine the chromosomal sex of the fetus,
the likelihood of Down Syndrome, etc.

An example that might hit closer to home these days is that RNA extraction is
used during many COVID-19 tests. A swab containing nasal cells is collected and
the nucleic acids are extracted to see if RNA from SARS-CoV-2 (the virus that
causes COVID-19) is present in the cells. If the viral RNA is present, it is
considered a “positive” COVID-19 test. If the viral RNA is not present, it is
considered a “negative” COVID-19 test.

a. (3 marks) Which of the following items could be useful reagents/equipment

when performing an RNA extraction during a COVID-19 test? Highlight or
check ALL that apply.

❑ Salt
❑ Sugar
❑ Dish soap
❑ Bleach
❑ Vinegar
❑ Flour
❑ Alcohol
❑ Filter
❑ Water
❑ Baking soda
❑ Baking powder

b. (1 mark) Multiple Choice – Which of the following would NOT occur when
extracting RNA during a COVID-19 test? Highlight your answer.

A. Addition of buffer solution followed by rigorous mixing

B. Breaking open of nasal cells using a detergent
C. Stabilizing nucleic acids using NaCl and bicarbonate
D. Collection of nucleic acids from the aqueous layer

PART 2 – DNA CONSTRUCTION

Introduction
Now that you have extracted the DNA, you might be surprised to learn that it is rather
goopy! In fact, you might be quite struck by how something so goopy can be the
genetic code for all existing life! Furthermore, where is the so-called “double helix” that
is so commonly used to describe DNA?
To understand where the genetic code is within the “goop”, view Figure 3.4 below.
Believe it or not, within the DNA extracted from the strawberry and cheek cells, there
are large structures called chromosomes (recall that there are 46 chromosomes per
human somatic cell). While we can’t see them with the naked eye, these chromosomes
could be viewed using a light microscope. What is important to know is that each
chromosome is actually made of 1 long strand of DNA that is tightly coiled together. If
you unravel this strand, you will see that the strand has a double helix shape, and that it
is within this double helix that nucleotides exist forming the genetic code. If you had
access to an electron microscope (a highly specialized microscope that is able to
visualize molecules at the nanometer level), you would be able to see this double helix.
To visualize this better, this section of the lab will be used to construct a model of DNA
using basic household ingredients.

Figure 3.4: “The chromosome” by Biosciences for Farming in Africa is

licensed under CC CY-NC-ND 2.0 Procedure
1. In this section, you will be building a model of DNA using a DNA code (see below).
This code is 6 letters long and consists of a combination of A (adenine), C
(cytosine), T (thymine) and G (guanine). Specifically, it is a portion of the DNA
sequence from the hemoglobin-β gene – a gene that when mutated causes sickle
cell anemia.

CAC CTC

2. To build your model, you will need the following:

Legend:

Part of the DNA Suggested Material Alternative Material

Deoxyribose
sugar Marshmallows - large, white

Phosphate Marshmallows - small, white

Base - A Marshmallows - small, orange

Base - C Marshmallows - small, yellow

Base - T Marshmallows - small, pink

Base - G Marshmallows - small, green

Bonds Toothpicks

NOTE 1: While marshmallows are the easiest (and most fun!) to work with, don’t
make a special trip out to get these items – SAFETY FIRST. Instead, it is OK to
substitute them for other things that you already have in your house (paper clips,
poof balls, colourful post-its, candies, etc.). The key thing is that the deoxyribose
sugar, phosphate and bases should look different from one another, and the bases
should have four different colour options. For example, you could use a paper clip
for the sugar, a poof ball for the phosphate and 4 different colours of M&Ms for the
bases. If you’re looking for an alternative for the bonds, consider using bobby pins,
twist ties, pieces of string, etc. - anything that helps show a bond or connection point
between the various objects.

NOTE 2: If you are using alternative parts to build your model, make sure to enter
them into the legend/chart above so that your instructor can accurately mark your
report.

3. You are now ready to begin building. Recall that

each letter in the code above actually represents a
nucleotide. The nucleotide is the monomer of
nucleic acid and contains 3 basic parts – a 5-
 carbon pentose sugar called deoxyribose, a
phosphate group, and one of four possible
nitrogenous bases (A, C, T or G). A and G are
purines and C and T are pyrimidines.
Begin by constructing 6 individual nucleotides, one
for each letter of the code given to you above. To
build these nucleotides, use the marshmallows and
toothpicks provided and follow the legend and
diagram provided.

4. Next, use toothpicks to form a polynucleotide with

all 6 nucleotides connected together. The
polynucleotide is the polymer of nucleic acid. The
nucleotides connect via a phosphodiester bond
between the phosphates and the deoxyribose
sugars. This forms the sugar-phosphate
backbone. The DNA sequence then refers to the
order of nitrogenous bases that stick out from the
sugar-phosphate backbone.

5. Next, make the polynucleotide double-stranded using complementary base

pairing. Use the legend and diagram provided.

Legend:
A pairs with T Connected via 2 hydrogen
bonds 2 toothpicks
C pairs with G Connected via 3 hydrogen
bonds 3 toothpicks
Other notes:
a. A purine always pairs with a
pyrimidine during complementary
base pairing
b. The first strand is “right side up”
and the second strand is “upside-
down.” As the 2 strands run in
 opposite directions, we say that
the DNA strands are anti-parallel.
c. At this stage, the DNA strand
resembles a ladder where the
nitrogenous bases form the
“rungs” of the ladder and the
sugar-phosphate backbone forms
the “poles” of the ladder.

6. Take a picture of your DNA strand and paste it into this document in the box below.
Use the following checklist and sample image (Figure 3.5) to make sure that all
graded elements are present.

❑ The sequence of your DNA strand matches

the DNA code given to you by your instructor
❑ All 12 nucleotides have 3 parts – phosphate,
deoxyribose sugar, and nitrogenous base
❑ The nucleotides within the sugar-phosphate
backbone are held together by
phosphodiester bonds
❑ The 2 polynucleotide strands are antiparallel
❑ Complementary base pairing is accurate – A
always bonds with T, C always bonds with G
❑ 2 hydrogen bonds are present between A’s
and T’s. 3 hydrogen bonds are present
between C’s and G’s.
❑ The double stranded DNA resembles a
ladder
❑ If alternative parts were used, the legend
was updated accordingly
(8 marks) Photo of your DNA model with the sequence CAC CTC: TOP
 BOTTOM

7. (4 marks) Examine the image below and identify the indicated parts of the DNA
strand in the spaces below:

a. Green box = __sugar phosphate backbone__

b. Red box = __nucleotide (Thymine)__
c. Yellow box = _pentose sugar_(deoxyribose)___
d. Blue arrow = ___double hydrogen bond___
e. Orange arrow = ___phosphodiester bond__

Figure 3.5 DNA model example

NOTE: Now that you’ve taken a picture of your “perfect” DNA strand, perform the next
2 steps keeping in mind that your structure will be destroyed during this process.**

8. Twist the 2 strands to form a double helix, similar to the DNA model found at your
bench. To do this, pick up the various edges of the DNA double stranded molecule
and attempt to twist it into a double helix shape. It may help to have an extra set of
hands. Do your best – it won’t be perfect.

9. Next, refer back to Figure 3.4. Remember that the double helix is actually coiled
tightly together to form a chromosome. While the largest human chromosome
contains ~250 million base pairs, for our purposes, form a chromosome using a 6-
nucleotide-long DNA strand.

a. To build a chromosome, mash your model all together into 1 big ball. Be
careful of the toothpicks! It is OK if a single tear falls down your cheek when
you do this. 
b. Consider now that the “goop” obtained from your DNA extraction is similar in
a way to the big ball of marshmallows. Even though it looks disorganized and
chaotic, it actually has a high level of organization hidden within.

10. (5 marks) If you had been asked to build a model of RNA instead of DNA, how would
your model look similar or different? To answer this question, list 2 similarities and 3
FULL differences in the boxes below:

DNA RNA
Difference 1 Double stranded Single Stranded

Difference 2 Base of Thymine Base of Uracil

Difference 3 Deoxyribose sugars Ribose sugars

Both DNA and RNA

Similarity 1 Both have sugar and phosphate backbones.

Similarity 2 Both use the bases Adenine, Guanine, and Cytosine.

PART 3 – KARYOTYPES

Introduction
Now that you have learned about DNA structure
and extraction, the final step in this lab is to view
various karyotypes under the microscope.
Karyotypes are defined as a complete set of
chromosomes within an organism. In a typical
human karyotype, 23 pairs of chromosomes are
extracted from a single nucleus, stained with a
dye, and arranged in order from largest to
smallest.

Using a light microscope, these chromosomes

can be observed at high magnification and used
to gather information about chromosomal Figure 3.6: “Normal male 46,XY human
karyotype” by Wessex Reg. Genetics Centre is
anomalies and cellular functions. licensed under CC BY 4.0)

For example, pregnant women who have chosen to undergo amniocentesis can
karyotype a fetal cell to find out the fetus’s biological sex or determine if missing
(monosomy) or extra (trisomy) copies of the chromosomes exist. A common
chromosomal anomaly is Trisomy 21 (Down Syndrome) in which a person has an extra
copy of the 21st chromosome. Karyotypes can also be used to make comparisons
between different species. For example, humans have 1 pair fewer chromosomes than
members of the great ape family (E.g. chimpanzees, bonobos, etc). Karyotypic
analysis suggests that this is due to an evolutionary event in which 2 ancestral
chromosomes fused together.
In this section, you will examine prepared human karyotypes and use this information
to draw genetic conclusions.
Procedure

1. Inspect the karyotypes below:

Karyotype A Karyotype B

Karyotype C Karyotype D

Karyotype E Karyotype F

2. The karyotypes A-F above can be analyzed in a variety of ways:

a. (2 marks) Indicate whether the following are chromosomally MALE or
FEMALE

Karyotype A: ___________MALE________________

Karyotype B: _________FEMALE__________________

b. (2 mark) Which karyotype(s) (A-F) were taken from a diploid SOMATIC cell?
List ALL that apply.

Karyotype(s): ____A, C, D, F_____________

c. (2 marks) Which karyotype(s) (A-F) were taken from a haploid SEX cell?
List ALL that apply.

Karyotype(s): _____B, E____________

d. (2 marks) TWO of the karyotypes above illustrate a TRISOMY. Indicate the

karyotypes (A-F) that illustrate this and what specific chromosomes (1-22,
X/Y) are affected.

Karyotype: _______C__________ Chromosome: ______21___________

Karyotype: _______ F__________ Chromosome: ____X/Y (has YY)___

e. (1 mark) ONE of the karyotypes above illustrates a MONOSOMY. Indicate

this karyotype (A-F) below and indicate what chromosome (1-22, X/Y) is
affected.

Karyotype: _____D____________ Chromosome:_ X/Y (only has one X)__

f. (1 mark) Fill-in-the-blank: A typical karyotype contains 22 pairs of

__autosomes__ and 1 pair of sex chromosomes.