Predicting Inheritance: Test Crosses

Predicting Inheritance: Test Crosses

 A test cross can be used to deduce the genotype of an unknown individual that is
expressing a dominant phenotype
 The individual in question is crossed with an individual that is expressing the
recessive phenotype
 This is because an individual with a recessive phenotype has a known genotype
 The resulting phenotypes of the offspring provides sufficient information to suggest
the genotype of the unknown individual

Results

 For a monohybrid test cross:

o If no offspring exhibit the recessive phenotype then the unknown genotype is
homozygous dominant
o If at least one of the offspring exhibit the recessive phenotype then the
unknown genotype is heterozygous
 For a dihybrid test cross:
o If no offspring exhibit the recessive phenotype for either gene then the
unknown genotype is homozygous dominant for both genes
o If at least one of the offspring exhibit the recessive phenotype for one gene but
not the other, then the unknown genotype is heterozygous for one gene and
homozygous dominant for the other
o If at least one of the offspring exhibit the recessive phenotype for both genes
then the unknown genotype is heterozygous for both genes

Worked example: Test crosses

 Rabbits have a single gene for ear length that has two alleles:
o D, a dominant allele that produces long ears
o d, a recessive allele that produces shorter ears
 A breeder has a rabbit called Floppy that has long ears and they want to know the
genotype of the rabbit
o There are two possibilities: DD or Dd
 The breeder crosses the long-eared rabbit with a short-eared rabbit
o A rabbit displaying the recessive short ear phenotype has to have the
genotype dd

Test cross possibility one table

Predicted ratio of phenotypes of offspring – 1 long ears

Predicted ratio of genotypes of offspring – 1 Dd

Test cross possibility two table

Predicted ratio of phenotypes of offspring – 1 long ears : 1 short ears

Predicted ratio of genotypes of offspring – 1 Dd : 1 dd

 The breeder identifies the different phenotypes present in the offspring

 There is at least one offspring with the short ear phenotype
 This tells the breeder that their rabbit Floppy has the genotype Dd
 If Floppy was genotype DD none of the offspring would have short ears

Exam Tip

Make sure before you start a test cross you think about the following: how many genes are
there, how many alleles of each gene are there, which is the dominant allele, what type of
dominance is it and is there linkage or epistasis between genes?
Predicting Inheritance: Chi-Squared Test
Predicting Inheritance: Chi-squared Test

 The difference between expected and observed results in experiments can be

statistically significant or insignificant (happened by chance)
 If the difference between results is statistically significant it can suggest that
something else is happening in the experiment that isn’t being accounted for
o For example linkage between genes
 A statistical test called the chi-squared test determines whether there is a significant
difference between the observed and expected results in an experiment
 The chi-squared test is completed when the data is categorical (data that can be
grouped)

Calculating chi-squared values

 Obtain the expected and observed results for the experiment

 Calculate the difference between each set of results
 Square each difference (as it is irrelevant whether the difference is positive or
negative)
 Divide each squared difference by the expected value and get a sum of these answers
to obtain the chi-squared value

Analysing chi-squared values

 To work out what the chi-squared value means, a table that relates chi-squared values
to probabilities is used
 If the chi-squared value represents a larger probability than the critical probability
then it can be stated that the differences between the expected and observed results are
due to chance
 If it represents a smaller probability than the critical probability then the differences
in results are significant and something else may be causing the differences
 To determine the critical probability biologists generally use a probability of 05 (they
allow that chance will cause five out of every 100 experiments to be different)
 The number of comparisons made must also be taken into account when determining
the critical probability. This is known as the degrees of freedom

Worked example: Chi-squared test

An experiment was carried out investigating the inheritance of two genes in rabbits; one for
coat colour and one for ear length. A dihybrid cross revealed the expected ratio of phenotypes
to be 9 : 3 : 3 : 1. Several rabbits with the heterozygous genotype were bred together and the
phenotypes of all the offspring were recorded. The ratio of the offspring was not exactly what
was predicted. In order to determine whether this was due to chance or for some other reason,
the chi-squared test was used.

Chi-squared worked example table

The expected ratio is calculated by multiplying the total number of organisms ( 128 rabbits)
with each expected ratio:

 In order to understand what this chi-squared value of 0.56 says about the data, a table
relating chi-squared values to probability is needed
 The chi-squared table displays the probabilities that the differences between expected
and observed are due to chance
 The degrees of freedom can be worked out from the results. It is calculated by
subtracting one from the number of classes
o In this example there are four phenotypes which means four classes, 4 – 1 = 3
o This means that the values in the third row are important for comparison
 For this experiment there is a critical probability of 0.05
o This means that 7.82 is the value used for comparison
 The chi-squared value from the results (0.56) is much smaller than 7.82
 56 would be located somewhere to the left-hand side of the table, representing a
probability much greater than 0.1
 This means that there is no significant difference between the expected and observed
results, any differences that do occur are due to chance

Genes, Proteins & Phenotype

 A gene can affect a phenotype of an organism
o A gene codes for a single protein
o The protein affects the phenotype through a particular mechanism
 The phenotype of an individual can also be affected by the environment

Phenotype Diagram
 Genes code for mRNA which gets translated into a polypeptide (protein) that affects the
phenotype of the organism

TYR gene & albinism

 Humans with albinism lack the pigment melanin in their skin, hair and eyes
 This causes them to have very pale skin, very pale hair and pale blue or pink irises in
the eyes
 There is a metabolic pathway for producing melanin:

1.
1. The amino acid tyrosine is converted to DOPA by the enzyme tyrosinase
2. DOPA is converted to dopaquinone again by the enzyme tyrosinase
3. Dopaquinone is converted to melanin

tyrosine → DOPA → dopaquinone → melanin

 A gene called TYR located on chromosome 11 codes for the enzyme tyrosinase
 There is a recessive allele for the gene TYR that causes a lack of enzyme tyrosinase or
the presence of inactive tyrosinase
 Without the tyrosinase enzyme tyrosine can not be converted into melanin

HBB gene & sickle cell anaemia

 Sickle cell anaemia is a condition that causes individuals to have frequent infections,
episodes of pain and anaemia
 Humans with sickle cell anaemia have abnormal haemoglobin in their red blood
cells
 β-globin is a polypeptide found in haemoglobin that is coded for by
the gene HBB which is found on chromosome 11
 There is an abnormal allele for the gene HBB which produces a slightly different
amino acid sequence to the normal allele
o The change of a single base in the DNA of the abnormal allele results in
an amino acid substitution
 The DNA base sequence GAG is replaced by GTG
 This means that CTC is replaced by CAC on the
complementary DNA template strand, meaning that GAG is
replaced by GUG in the resulting mRNA
 o This change in amino acid sequence results in an abnormal β-
globin polypeptide
 The amino acid Glu is replaced with Val
 The abnormal β-globin in haemoglobin affects the structure and shape of the red
blood cells
o They are pulled into a half moon shape
o They are unable to transport oxygen around the body
o They stick to each other and clump together blocking capillaries
 A homozygous individual that has two abnormal alleles for the HBB gene
produces only sickle cell haemoglobin
o They have sickle cell anaemia and suffer from the associated symptoms
 A heterozygous individual that has one normal allele and one abnormal allele for
the HBB gene will produce some normal haemoglobin and some sickle cell
haemoglobin
o They are a carrier of the allele
o They may have no symptoms

Sickle Cell Anaemia Diagram

Normal red blood cells and Sickle cell blood cells - The sickle cells cause a blockage in the
capillary and restrict blood flow

F8 gene & haemophilia

 Factor VIII is a coagulating agent that plays an essential role in blood clotting
 The gene F8 codes for the Factor VIII protein
 There are abnormal alleles of the F8 gene that result in:
o Production of abnormal forms of factor VIII
o Less production of normal factor VIII
o No production of factor VIII
 A lack of normal factor VIII prevents normal blood clotting and causes the
condition haemophilia
 The F8 gene is located on the X chromosome
 o This means F8 is a sex-linked gene
o Haemophilia is a sex-linked condition
 If males have an abnormal allele they will have the condition as they
have only one copy of the gene
 Females can be heterozygous for the F8 gene and not suffer from the
condition but act as a carrier

HTT gene & Huntington's disease

 Huntington’s disease is a genetic condition that develops as a person ages

 Usually a person with the disease will not show symptoms until they are 30 years old
or older
 An individual with the condition experiences neurological degeneration; they lose
their ability to walk, talk and think
 The disease is ultimately fatal
 It has been found that individuals with Huntington's disease have abnormal alleles of
the HTT gene
o The HTT gene codes for the protein huntingtin which is involved in neuronal
development
o People that have a large number (>40) of repeated CAG triplets present in
the nucleotide sequence of their HTT gene suffer from the disease
 The abnormal allele is dominant over the normal allele
o If an individual has one abnormal allele present they will suffer from the
disease

Gene, protein & phenotype summary table

Key molecules
Gene Genotype Phenotype
involved

Tyrosinase enzyme Homozygous for

TYR Albinism
and melanin abnormal allele
Heterozygous Carrier
HBB Haemoglobin Homozygous for
Sickle cell anaemia
abnormal allele
Heterozygous female Carrier
F8 Factor VIII Homozygous for
Haemophilia
abnormal allele

Heterozygous Huntington's disease

HTT Huntingtin
Homozygous for
Huntington's disease
abnormal allele
Exam Tip

You may be asked to predict the inheritance of diseases like the ones above.
An example question would be: Max and Jane are trying for a baby but they are concerned
about the possibility of their child having haemophilia. Neither Max or Jane have
haemophilia themselves but Jane’s father had the condition. What are chances that their
child could have haemophilia?

For questions like this, it is very important to gather early on whether the abnormal allele that
causes the disease is dominant or recessive and if there is any sex linkage. In this example
for haemophilia, the abnormal allele is recessive and the gene is sex-linked. Then the next
step would be to work out the genotypes of the parents from the information given and use
this to create a genetic diagram.