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Biology 156: An Introduction to Mendelian Genetics
Dr. Cheryl Blake: Based on the lab from the Desert Vista Lab
Manual
There are no special safety considerations associated with this lab.
BACKGROUND: AN INTRODUCTION TO MENDELIAN GENETICS
Gregor Mendel, an Austrian monk, was the first we know of who
completed and documented a scientific study of the progeny of controlled
matings. His work was unique in that he carefully controlled the inputs (the
types of plants that he was using), replicated his work hundreds of times to
ensure the data were not due to random variation, and recorded the outcomes
of much of his work. The process that he followed is shown in the picture
below:
Mendel’s choice of the pea plant for his
work was advantageous. Not only is this
an organism that has a fast reproductive
cycle, and one that generates many
offspring in a very short period of time,
but the pea plant's flowers have useful
qualities as well. Left alone, the flower
will self-pollinate (see sec. 9.2B), but
these flowers can easily be crosspollinated as well as you see in the
picture.
These are the two key options that a
geneticist would like to have for
organisms. Other famous geneticists have
worked with the fruit fly Drosophilia
melanogaster and the nematode worm
Caenorhabditis elegans (C. elegans),
which have many of the same favorable
qualities. Imagine doing genetic crosses
with elephants: not only would it take
years to produce one offspring, but you
would have to feed them too!
In this lab, we will work with the organism maize (corn) which was
favored by the famous geneticist Barbara McClintock – the discoverer of
“jumping genes”. Corncobs are the fruit from one corn plant. Each seed is
the product of a cross between the “mother” plant on which the cob was
formed and a pollen grain carrying sperm from a “father” plant. An
advantage of corn is that the many progeny – the seeds – are aligned for easy
data collection.
Mendel began his study of genetics without knowledge of cell structure,
mechanisms of cell division, DNA, proteins and other key biological ideas
that we take for granted. Some of his contemporaries were describing the
cellular nature of life (Schleiden & Schwann and Virchow) while another
famous biologist – Charles Darwin – was describing how natural selection
could explain the patterns of evolution he had observed. You will begin this
lab as Mendel did by observing the outcomes of crosses. You will then
develop an understanding of why these outcomes are as they are.
THE NATURE OF THE GENETIC CROSS
A hybrid is the result of the breeding of two individuals. In a
monohybrid cross like the one pictured on the previous page, one trait of
the organism is the focus of the study (the color of the flowers in this
case);
other traits are simply ignored. Examining the eye color of the offspring
that
results from a mating between a blue-eyed person and a brown-eyed person
is an example of a monohybrid cross. The parents and offspring clearly
have other features such as height, hair color, intelligence etc., but these
traits are ignored. Similarly, a dihybrid cross occurs when the experimenter
attends to two traits (e.g., eye color and height) in the parents and
offspring.
One of the unique features of Mendel’s work was that he always carried
his crosses through two generations; you will see why this was so critical as
the lab progresses. To simplify recording of information in genetic crosses,
the generations are named. The original parents are the P generation. Their
offspring comprise the F1 (first filial) generation. Mendel always allowed
offspring in the F1 generation to mate with each other; the progeny from this
second generation (the “grandchildren” of the original parents) comprise the
F2 (second filial) generation.
Another unique feature of Mendel’s crosses was that he began all of his
work with true-breeding strains for the trait he intended to study. A truebreeding or purebred organism is one that produces identical offspring
generation after generation when mated to an organism of the same breed
(e.g., a purebred collie mated with another purebred collie will have pups
that all have collie characteristics). Sec. 9.2D in you text shows diagrams
of
seven of the traits that Mendel studied as he mated his pea plants. He had,
for example, purebred strains of pea plants that always produced purple
flowers and strains that always produced white flowers at his disposal. In
today’s activity, we will attend to two traits found in corn kernels: the
color
of the kernel (purple or yellow) and the amount of starch found in the
kernel.
Starchy kernels are plumper, while sweeter kernels appear to be shriveled in
comparison.
EXERCISE A: DETERMINING DOMINANCE & PHENOTYPIC RATIOS IN A
MONOHYBRID CROSS
(a) Obtain a bag of corncobs. A different genetic cross has produced each
cob.
(a) Take out the corncobs labeled A (all purple kernels) and B (all yellow
kernels). Imagine that seeds from each were planted and pollen (sperm)
from the plants growing from the yellow seeds were mated with the eggs
from the plants that grew from the purple seeds. A cob from this cross
would contain many seeds. These seeds would be the
generation, while the original purple and yellow seeds would comprise
the
generation.
(b) The cross described in question 1 was actually done by a biological
supply company. The offspring are present on cob C. Describe these
offspring.
(c) What are some hypotheses that might explain the appearance of cob C?
(Try to think like Mendel: pretend that you don’t know about cells, cell
division, DNA or genetics!)
(d) The same biological company allowed kernels like those on cob C to
grow into mature plants, and allowed them to pollinate each other. Cob
D, the
generation, clearly consists of both purple and yellow
kernels. Did this second cross disprove any of your hypotheses from
question 3? Explain.
(e) Working with your partner, identify the color of 100 kernels on your D
cob. Record the number of your purple and yellow kernels here:
Purple
Yellow
(f) Add the data for all the D cobs in the class below:
Purple
Yellow
Total
(g) What was the ratio of purple to yellow kernels for the class as a whole?
In other words, what percentage of the kernels in this second generation
were purple and what percentage were yellow?
%Purple (# purple/total)
%Yellow
(#
purple/total)
Ratio of purple to yellow (divide the % of purple by the % of yellow)
(h) Complete the data table* of Mendel’s work below. Then comment on
the similarities and differences within his data and between these data
and yours.
P trait A
X
P trait B
# trait A
in F2
5474
# trait B in
F2
1850
Total
F2
7324
Ratio in F2
Spherical
Wrinkled
2.96 : 1
seeds
Yellow seeds
Green
6022
2001
Purple
White
705
224
flowers
Inflated pods
Constricted
882
299
Green pods
Yellow
428
152
Axial
Terminal
651
207
flowers
Tall stems
Dwarf
787
277
*Adapted from Table 10.1 in Purves et. al., LIFE, The Science of Biology
5th Ed., 1998
BACKGROUND CONT.: THE PUNNETT SQUARE - PREDICTING OUTCOMES IN
MONOHYBRID CROSSES
Now, consider what you know about genetics, cell structure and cell
division. As you studied the process of meiosis in the previous lab, you
learned that the daughter cells from a meiotic division have half the number
of chromosomes found in the parent cell. You also learned about the nature
of the gene: genes, of course, are simply the directions for making cellular
proteins. Finally, you learned that homologous chromosomes have the
same genes in the same order, but the specific directions on the two
homologous chromosomes might vary.
The genetics term "allele" refers
to the possible versions that a
particular gene might have. The
gene locus is the position on a
particular chromosome in which
we find the gene. In this figure,
we see a pair of homologous
chromosomes. The locus for
flower color is identified, and
two versions of the gene - the
allele for purple flowers and the
allele for white flowers are
shown.
In our corn kernels, there are two alleles at a particular locus that
controls
the color of the kernel. One allele codes for an enzyme (a protein) that
catalyzes the production of a purple pigment. The alternative allele at
that locus does not code for a functional enzyme, and kernels are the
“default” color yellow (i.e., they are “purple lacking”).
When Mendel crossed pea plants with
purple flowers with purebred plants with
white flowers, all the offspring in the
first generation had purple flowers.
Similarly, the F1 generation seeds
(peas) resulting from a cross of tall and
short plants were all tall. In all cases
that Mendel recorded, one trait appeared
to overpower the other trait in the F1
generation. He termed this trait the
dominant trait. Interestingly, the trait
that had been masked – the recessive
trait – reappeared in the second
generation (F2) as you see in the figure.
Mendel’s careful work over two generations demonstrated that the recessive
trait had not truly been "overcome"; it was not gone, but simply being
masked in the first generation. His identification of a dominant trait in the
F1 generation also showed that, contrary to popular belief, these traits did
not simply blend to produce a hybrid mixture. For example, purpleflowering X
white-flowering plants did not produce plants of medium or
blended color - they were all purple; tall plants X short plants were not of
medium height - they were all tall.
Mendel described a particulate hypothesis to explain his observations.
He suggested that each parent had two particles that provided the
information for a specific trait, and that one of each of these particles
must
be present in the egg or sperm produced by the parent. Purebred parents
would have only one type of particle (but two copies). Mendel developed
the habit of using a capital letter to identify the dominant trait. He used
the small letter to identify the alternative, recessive trait. For example,
once he had identified that tall trait dominated the short trait in the F1
generation, he identified the tall-producing particle (remember that he did
not yet know what genes were) as “T” while short-producing particles were
assigned “t”. Thus a purebred tall parent would have two T particles, while
a purebred short parent would have two t particles. The tall parent would
make eggs (or sperm) with one T particle, while the short parents sperm (or
eggs) would contain one t particle. Fertilization would result in an
offspring
with one of each particle (one T and one t), and the observed, dominant, tall
appearance.
(i) Which trait is dominant for kernel color in corn?
Examine the other cobs in your bag (you’ll need the key to identify
which is which) and determine which trait is dominant: sweet (shriveled)
or starchy (plump).
Return your attention to the structure of a chromosome and the concept
of a gene. We know that each cell has two homologous chromosomes with
loci for many traits. The diagram below shows three possibilities for
chromosome combinations in a situation such as you saw corn. If you work
as Mendel did, your identification of purple as the dominant trait in the
first
exercise should lead you to identify the purple allele as “P” with the yellow
allele as “p”.
(j) Fill in the three possibilities for allele combinations that could be
present
in kernels below. Then identify whether the appearance of the kernel
would be purple or yellow given that genetic content. The first
possibility is done for you to serve as a model.
P
P
Purple kernel
_________
The pair of alleles that are present at a
particular locus is the organism’s genotype
(PP, Pp or pp). Its appearance or a
functional quality it possesses is its
phenotype (purple or white for these pea
plants). When the alleles are the same, the
genotype is homozygous (PP or pp); when
they are different, it is said to be
heterozygous (Pp).
The allele combinations you
identified on the previous page
should be PP, Pp and pp; these are
the three possible genotypes that
can occur for this trait in corn. Since
there are two ways to be
homozygous (PP or pp, in this case),
the term dominant or recessive is
added for clarity. Thus PP is the
homozygous dominant genotype,
and plants with this genotype have
the purple phenotype due to the
presence of the dominant allele (i.e.,
they have the ability to make the
enzyme needed to produce the purple
pigment).
Plants
with
the
heterozygous genotype (Pp) are
purple as well, since they also
produce the necessary enzyme, but
only those with the homozygous
recessive genotype (pp) have the
yellow phenotype since only these do
not have the enzyme and thus retain
their “natural” color.
The phenotypes that Mendel described were all physical appearances
observable with the naked eye. Our advanced technology allows us to
identify phenotypes that are biochemical and cellular in nature, such as
whether a person has a disorder such as sickle-cell anemia. In this case, the
phenotype is really a set of symptoms that include shortness of breath and
joint pain during activity. Since this disorder is controlled by a single
gene
locus, a person can have the genotypes SS, Ss or ss; only with the ss
genotype do the phenotypic symptoms appear. Many of the symptoms of
sickle cell are observable only with a microscope or by using biochemical
techniques.
Using your knowledge of meiosis and the vocabulary that you now know,
you can effectively and easily predict the genoptypic and phenotypic
outcome of genetics crosses by using Punnet squares. First, you must
determine whether the trait you are studying adheres to the
dominant/recessive patterns described by Mendel.
(k) How would you determine whether the height of a corn plant followed
the tall vs. short pattern described by Mendel? If it did, how would you
know which trait was dominant?
Once the genetics pattern for a particular trait has been established, and
the genotypes of the parents are known, you can create a Punnett square to
predict the types of offspring that would be produced from that cross. To
complete a Punnett square, first determine the possible alleles that would be
present in the sperm or egg produced by each parent. A tall pea plant that
has a homozygous genotype (i.e., one that is TT) can only produce eggs or
sperm that carry the T allele. Convince yourself that this is true by
reviewing how meiosis produces gametes: the available DNA (in this case,
containing the T allele) is copied and then sorted over two divisions into
gametes. Since only the T allele is present to be copied during DNA
replication, only the T allele can be present in the gametes. If the tall
parent
were heterozygous (Tt), 50% of the gametes would carry the one T allele,
while 50% would contain a single t allele for that locus.
The next step is to place the allele possibilities for that parent’s gametes
along one axis of the square. Repeat this procedure for the other parent. It
is important to note that a Punnet square may not have the same number of
gamete options on the vertical and horizontal axes. (The example below
demonstrates this fact.) Complete the square by modeling fertilization
events: combine the gametes from each parent in the center boxes.
Determine the phenotype of each child using the information regarding
dominance that you have already determined. Finally, identify the
genotypic and phenotypic ratios in this new generation.
An example of a Punnet square completed for a cross of a heterozygous
tall pea plant with a short pea plant is shown on the next page. The logic
and steps you would follow to determine the genotypic and phenotypic ratios
of the plants produced from this mating follow:
1. We first must determine the genotypes of both parents. We already know
from Mendel’s work that tall is the dominant trait, so we assign the letter
T for tallness and t for shortness. The heterozygous parent must be Tt.
The genotype of the short parent is not given, but to express the
recessive genotype, it must be tt.
2. One cell in the tall parent (Tt) undergoing meiosis would produce 4
gametes: two would carry the T allele, while 2 would carry the t allele.
In other words, half of the haploid gametes produced by this diploid cell
would be T, and half would be t. This information is entered along the
vertical axis of the square below.
3. The short parent (tt) can only produce haploid gametes carrying the t
allele. This single possibility is listed on the horizontal axis as shown.
4. The possibilities are combined in the central areas by simply writing
both the alleles from the vertical and horizontal axis bordering that box.
The order of the alleles is irrelevant, but convention provides for writing
the dominant allele, if it exists, first.
5. The phenotype of each offspring is determined: if at least one T is
present, the organism is tall, while only those with a homozygous
recessive genotype are short.
6. The genotypic and phenotypic ratios are calculated. In this case, they
happen to be identical.
Tall, heterozygous parent (Tt)
X
Short parent (tt)
Produces half T and half t gametes
produces only gametes with t
Tall parent’s gametes
T
t
Short parent’s gamete
t
Tt – tall offspring
tt – short offspring
The genotypic ratio in the offspring of this cross is 1 Tt : 1tt (50% to
50%)
The phenotypic ratio in the offspring is also 1 tall : 1 short
Note: you could create a 2 X 2 square (the more traditional method) for
this cross as shown below. However, since the short parent’s gametes
would be alike in both columns, the end result is identical. Use a 2 X 2 box
like this one (as the computer does in the exercise below), if this is easier
for
you.
Short parent’s gametes
t
Tt – tall offspring
tt – short offspring
t
Tt – tall offspring
Tall parent’s gametes T
t
tt
–
short
offspring
The genotypic ratio in the offspring of this cross is 1 Tt : 1tt (50% to
50%)
The phenotypic ratio in the offspring is also 1 tall : 1 short
EXERCISE B: USING PUNNETT SQUARES TO DETERMINE THE OFFSPRING
IN A MONOHYBRID CROSS
7. At this point, examine only one characteristic to complete only
monohybrid crosses.
(We will examine dihybrid crosses later.)
Remember that Mendel always began his work with pure breeding
parents, and carried his crosses over two generations. In this case, you
can begin with any parents, but the computer only allows you to directly
examine the F1 generation. By using the offspring produced in this
generation as the parents for the next, however, you can model a
twogeneration monohybrid cross like those Mendel did.
BACKGROUND: THE TEST CROSS
A test cross is a unique type of cross used for only one purpose: to
determine the genotype of an organism that displays the dominant
phenotype but whose genotype is unknown. Imagine, for example, that you
found a pea plant that had purple flowers in your garden and that you knew
nothing about its "parents". You know only that purple flowers are dominant
to white flowers, and you would like to know what genotype this plant has.
Mendel devised a simple system for
answering this question that works for
many organisms. He simply mated this
plant with purple flowers with a pea
plant that produced only white flowers
(i.e., it had the recessive phenotype).
He then examined the offspring. If
many offspring were produced, and all
the offspring peas were purple, the
probability that the parent purple plant
had the PP genotype was very high. If,
however, this mating produced some
white flowers, the parent must have
been Pp. Consider why this is so, and
why we speak of a high probability in
the first case and a certainty in the
second case by following the logic on
the next page:
1. Since "purpleness" is dominant, the purple parent can be either PP or
Pp. The parent with white flowers chosen for mating must have the pp
(homozygous recessive) genotype (otherwise it wouldn't have white
flowers), so this information is known.
2. IF the purple parent was PP, it would produce only P-containing
gametes. The white pea plant can only produce p-containing gametes.
Fusing these two gametes would result in plants with the Pp genotype,
which would be purple.
3. IF the round parent were Pp, half of its gametes would contain the P
allele, while the other half would carry the p allele. As the Punnett
square on the previous page shows, half of these offspring would be
purple and half white.
4. Thus, it is the offspring that tell us about the parent. IF there are many
offspring, and IF all of these offspring have purple flowers, then, as
stated above, the probability that the parent was PP is quite high. To
understand why we are not certain, consider that this is a case of
probability. It is possible, for example, that we could flip a coin 10 times
and end up with 10 heads and no tails. This does not mean that the coin
does not contain a “tail side”, necessarily; it might have both a head and a
tail, but random chance might result in the heads-only result. The greater
the number of flips that result in only heads, however, the greater our
belief becomes that no tail side is to be found on this coin. 1000 heads in
1000 flips would afford us great confidence that this coin does not have a
tail side. Similarly, 1000 round offspring would be much more
convincing of the lack of an r allele than 5 would be.
5. Consider why, on the other hand, even a single white-flowered offspring
convinces us that, unless a mutation has occurred, the parent must have
been Pp. Any white offspring expressing the recessive genotype must
have the pp genotype. Since offspring must receive alleles from each
parent, the purple parent must have had one p to give its progeny. Since
it had purple flowers, it must have had one P allele as well, so its
genotype must be Pp.
EXERCISE C: EXAMINING THE RESULTS OF A TEST CROSS IN CORN
8. The biological supply company has also completed a test cross of a
purple kernel (a corn seed) of unknown origin. As is done in all test
crosses, they mated a plant grown from this kernel, with a plant
expressing the recessive phenotype. The corn plant with the recessive
phenotype would have the genotype
(color)
and the phenotype
. The progeny of this cross are on cob E. First count
enough kernels to effectively describe the phenotype or phenotypic ratio
on this cob. (You are free to combine your data with other groups as you
did in the first exercise to increase your sample size.) Record your data,
and explain what these observations tell you about the original purple
kernel. Make sure that the logic you present in your conclusion is
thorough and clear.
BACKGROUND: THE DIHYBRID CROSS
In a dihybrid cross, two traits or characteristics of parents and offspring
are attended to. In Mendel’s case, he might have attended both to the height
of a pea plant and the color of its flowers while ignoring characteristics of
its
pods and seeds. Since two traits are being examined, the genotypes at two
loci must be considered. For example, a tall plant with purple flowers that
is homozygous for both traits would have the genotype TTPP, while one
that is also homozygous for its height but is heterozygous for its flower
color
would be TTPp. A short plant with white flowers must be ttpp.
Though the process of creating a Punnett square is the same, determining
the alleles in the gametes can be a little more difficult. The important
thing
to remember is that all gametes must contain one allele for each trait in the
organism. In the above example, each gamete should have an allele for
height and an allele for flower color. For example, gametes produced from
the TTPp parent might be TP or Tp.
9. Why couldn’t this plant produce a gamete with the tp genotype?
As he did with the monohybrid cross, Mendel began with purebred
parents and examined the offspring in the F1 generation. He allowed these
offspring to self-pollinate, and then recorded the phenotypes in the F2. Our
understanding of meiosis and Punnett squares allows us to explain the
9:3:3:1 phenotypic ratio that Mendel observed in these dihybrid crosses.
Follow the argument presented in section 9.5 and examine the figure below.
In this figure, Mendel attended to the color and the shape of the seeds. The
data demonstrate that the alleles for the two traits (color - Y or y, and
shape R or r) assort independently; in other words, the off spring can have
any
combination of traits as long as the original alleles were present in their
parents. (Note: this is not true if the alleles are close together on the
same
chromosome.) When you understand the figure below, complete the
remaining exercises to become familiar with the dihybrid cross
EXERCISE E: EXAMINING THE RESULTS OF A DIHYBRID CROSS IN CORN
12.Recall that for kernel color, purple is dominant to yellow. The presence
of a gene that codes for an enzyme that catalyzes the synthesis of sugar
molecules into long starch polymers makes the kernels plump with starch
granules.
Kernels that lack this gene are sweeter and have a more
shriveled appearance (the recessive phenotype).
10.In a Mendelian dihybrid cross, one parental type is homozygous
dominant at both loci; its genotype at these two loci is
. The other parental type is homozygous recessive; its genotype is
. Explain why all F1 offspring will be heterozygous at both
loci.
11.The genotype of the F1 plants is __________. Each of these F1 plants
can make four types of gametes. They are:
12.Make a Punnett square of an F1 x F1 dihybrid cross. Diagram this
cross here. Once the interior of the square contains the genotypes of each
of these F2 offspring, determine the phenotype of each F2 organism.
Finally, determine the expected phenotypic ratio in the F2 by counting
the number of each phenotype. Record this information and circle it.
13.Take out a cob that is a result of a dihybrid cross from your bag. Score
several rows of kernels on the F2 cob for kernel shape and color. (Sample
at least 100 kernels; remember that it is fine to combine your data with
other groups to increase your end sample size and thus improve your
confidence in your data.) Record your results in the chart below. Also
indicate the relative proportions of each phenotype by dividing the
smallest number into each of the larger ones.
Kernel Shape and Color on F2 Cob.
Phenotype
Total
Phenotypic Ratio
Starchy, Purple
14.Compare the relative proportions of each of the four phenotypes that you
observed with those that you expected from your Punnett square results.
How similar were your results to those that you had predicted?
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