Types of genetic tests
1. Cytogenetic
2. DNA
3. Metabolic
Karyotype
• Picture of the chromosomes in a cell used to check for
abnormalities
Prenatal diagnosis: trisomy 21 (Down’s syndrome)
Postnatal diagnosis:
Postnatal diagnosis: detecting cancer
Preparing a karyotype
• Harvest cells (from where?)
Postnatal diagnostic karyotype
Prenatal diagnostic karyotype
Preparing a karyotype
• Harvest cells
Postnatal diagnostic karyotype
tumor biopsy
skin cells from mouth (ie for non-cancer related diagnoses)
Prenatal diagnostic karyotype
chorionic villi sampling (CVS)
amniocentesis
Who is offered amniocentesis or CVS?
• Maternal age (women 35 or older)
Risk of Down’s syndrome:
mother in 20s 1/1250
99.92% OK
mother at 35 1/400
99.75% OK
mother at 40 1/100
99% OK
Down’s syndrome: how does it happen?
Chromosomal non-disjunction during meiosis of eggs
and sperm.
1. Chromosomes
replicate
2. Homologous
chromosomes
separate
3. Chromatids
from each
chromosome
separate
Who is offered amniocentesis or CVS?
• Maternal age (women 35 or older)
Risk of Down’s syndrome:
mother in 20s 1/1250
99.92% OK
mother at 35 1/400
99.75% OK
mother at 40 1/100
99% OK
• A previous child or pregnancy with a birth defect
• Screening test with a positive result
• Other family history
Prenatal diagnosis:
amniocentesis
• Sampling cells from amniotic fluid
• Usually done ~ 15–18 weeks
Prenatal diagnosis: chorionic villi sampling (CVS)
• Sampling cells from placenta
• Usually done 10–12 weeks
Preparing a karyotype
• Harvest cells
• Culture cells 1–2 days
• Arrest cells in metaphase with colchicine
metaphase
chromosomes
condense
Mitosis
DNA
replication
nuclear
envelope
breaks
down
metaphase
chromosomes
aligned on
spindle fibres
Preparing a karyotype
• Harvest cells
• Culture cells 1–2 days
• Arrest cells in metaphase with colchicine
• ‘Spread’ cells on slide and
stain
• Count chromosomes in 20
representative cells
• Capture image of five ‘best’
cells and construct
karyotypes for each
metaphase
FISH analysis of chromosomes:
Fluorescent In Situ Hybridization
Metaphase spread
chromosomes
stained with DAPI, a
fluorescing stain that
specifically binds
double-stranded
DNA
FISH
Expose DAPI-stained
metaphase chromosomes to
fluorescent probes
red = control probe for
centromere of the X
chromosome and another
probe for end of chromosome
X
green = probe for the end of
chromosome 4
DiGeorge syndrome/CATCH22
• Microdeletion on chromosome 22
• Birth defect that affects the immune system
• Absence or underdevelopment of the thymus and
parathyroid glands
• Facial features include low-set ears, wide-set
eyes, small jaw and bowing up of upper lip
FISH tests: DiGeorge syndrome
Expose DAPI-stained
chromosomes to
mixture of fluorescent
probes
green = control probe
for chromosome 22
red = probe for
DiGeorge region on
long arm of
chromosome 22
FISH tests: Painting chromosomes
Expose chromosomes to fluorescent probes that highlight
entire chromosomes.
FISH tests: Painting chromosomes
Expose chromosomes to fluorescent probes that highlight
chromosomes 13, 18, 21, X and Y.
nuclei from
the same
foetus
green = chromosome 13
red = chromosome 21
aqua = chromosome 18
green = X chromosome
red = Y chromosome
Trait
A physical characteristic that is determined
by genes, eg eye colour.
Human traits
Thumb shape
hh
Hh or HH
‘Hitchhiker’s thumb’
Earlobe attachment
AA or Aa
aa
unattached
attached
Human traits
Thumb shape
hh
Hh or HH
‘Hitchhiker’s thumb’
Genotype vs phenotype
Genotype = specific allelic make-up of an individual, eg HH, Hh or hh
Phenotype = an observable physical or measurable biochemical
characteristic, eg thumb shape or lack of a particular enzyme
Punnett squares
Remember these??
Used to determine the probability
of an offspring having a particular
genotype
H
H
h
H
hh
‘Hitchhiker’s thumb’
Hh or HH
H allele = dominant
h allele = recessive
Punnett squares
Now try it backwards
hh
‘Hitchhiker’s thumb’
Hh
Hh
hh
hh
Hh or HH
H allele = dominant
h allele = recessive
Recombination: Shuffling the deck
DNA crossovers in chromosome pairs that result in children
receiving a different combination of genes than either parent