Cytogenetics Lesson 3

8/23/2019 Cytogenetics Lesson 3

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Cytogenetics: Karyotypes and Chromosome

Aberrations

Cytogenetics = The study of chromosome number,

structure, function, and behavior in relation to gene

inheritance, organization and expression

Chromosome Chromo = colored in response to dye Some = body Chromosome of Eukaryotes have been the traditional

subject for cytogenetic analysis because they are largeenough to be examined with light microscope


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Chromosome Number

Chromosome number in selected organisms


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Cytogenetic methods to detect chromosomal

abnormalities underlying human birth defects usually

involve analysis ofmitotic chromosomes


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Why Analyse Chromosomes and Genes?

Genetic errors arise from deletions orinsertions ofgenetic material, abnormal numbers of whole

chromosomes or genes, and even from

misplacement ofa single base in the DNA

sequence.

Genetic abnormalities can range from relatively

harmless to severe: from vitamin deficiencies and

food allergies to cancer, birth defects and infant

mortality.


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Chromosome Shape

As chromosomes condense and become visibleduring cell division, certain structural features can

be recognized

CentromereA region of a chromosome to which microtubule

fibers attach during cell division

The location of a centromere gives a chromosome itscharacteristic shape


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Centromere Location

MetacentricA chromosome that has a centrally placed

centromere

SubmetacentricA chromosome whose centromere is placed closer to

one end than the other

AcrocentricA chromosome whose centromere is placed very

close to, but not at, one end


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Human Chromosomes

Replicated chromosomes at metaphase consist ofsister chromatids joined by a single centromere

Fig. 6-1, p. 122


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Chromosome

Sister Chromatids


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Metaphase Chromosomes

Chromosomes are identified by size, centromerelocation, banding pattern


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Metacentric Submetacentric Acrocentric

Short

arm(p)

Satellite

p Centromere

p

Stalk

Long

arm(q)

q q

3 17 21


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Types of Chromosomes

Sex chromosomes In humans, the X and Y chromosomes that are

involved in sex determination. These have different

sizes and shapes

Autosomes Chromosomes other than the sex chromosomes In humans, chromosomes 1 to 22 are autosomes


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A Set of Human Chromosomes

Human chromosomes are analyzed by constructionofkaryotypes

KaryotypeA complete set of chromosomes from a cell that has

been photographed during cell division at the

metaphase stage and arranged in a standard

sequence


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A Human Karyotype


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Karyogram:

Chromosome Banding Patterns


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System of Naming Chromosome Bands

Allows any region to beidentified by a descriptive

address (chromosome

number, arm, region, and

band)


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Add a fewdrops of blood.

Add phytohemagglutininto stimulate mitosis.

Draw 10 to 20 ml

of blood. Incubate at 37Cfor 2 to 3 days.

Transfer to tubecontaining fixative.

Transfercells to tube.

Add Colcemid toculture for 1 to 2

hours to stop mitosisin metaphase.

Centrifuge toconcentrate cells. Addlow-salt solution toeliminate redblood cells andswell lymphocytes.

Drop cells ontomicroscope slide.

Examine withmicroscope.

Digitizedchromosomeimages processedto make karyotype.

Stain slidewith Giemsa.


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Metaphase Chromosomes (a)

Arranged Into a Karyotype (b)


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Constructing and

Analyzing Karyotypes

Karyotype construction and analysis are used toidentify chromosome abnormalities

Different stains and dyes produce banding patternsspecific to each chromosome

Karyotypes reveal variations in chromosomalstructure and number

1959: Discovery that Down syndrome is caused byan extra copy of chromosome 21

Chromosome banding and other techniques canidentify small changes in chromosomal structure


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Information Obtained from a Karyotype

Number of chromosomes Sex chromosome content Presence or absence of individual chromosomes Nature and extent of large structural abnormalities


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Four Common Chromosome Staining

Procedures


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Banding technique Appearance of chromosomes

G-banding Treat metaphase

spreads with trypsin, an enzyme that

digests part of chromosomal protein.Stain with Giemsa stain. Observebanding pattern with light

microscope.

Darkly stained G bands.


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Q-banding Treat metaphase

spreads with the chemical

quinacrine mustard. Observefluorescent banding pattern with aspecial ultraviolet light microscope.

Bright fluorescent bands

upon exposure to

ultraviolet light; same asdarkly stained G bands.



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R-banding Heat metaphase

spreads at high temperatures to

achieve partial denaturation of DNA.Stain with Giemsa stain. Observewith light microscope.

Darkly stained R bands

correspond to light bands

in G-banded chromosomes.Pattern is the reverse of G-banding.



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C-banding Chemically treat

metaphase spreads to extract DNAfrom the arms but not the

centromeric regions ofchromosomes. Stain with Giemsa

stain and observe with light

microscope.

Darkly stained C band

centromeric region of thechromosome corresponds

to region of constitutiveheterochromatin.



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Chromosomal Aberrations and Specific

Syndromes


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Chromosome Painting

New techniques using fluorescent dyes generateunique patterns for each chromosome


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Obtaining Cells for Chromosome Studies

Any nucleus can be used to make karyotype Lymphocytes, skin cells, cells from biopsies, tumor

cells

Sampling cells before birthAmniocentesis Chorionic villus sampling (CVS) Cord Blood


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Amniocentesis

A method of sampling the fluid surrounding thedeveloping fetus by inserting a hollow needle and

withdrawing suspended fetal cells and fluid

Used in diagnosing fetal genetic and developmentaldisorders

Usually performed in the sixteenth week ofpregnancy


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Amniocentesis


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Removal of about 20 ml of amniotic fluid containing

suspended cells that were sloughed off from the fetus

A few biochemicalanalyses with some

of the amniotic fluid

Centrifugation

Quick determination of

fetal sex and analysis

of purified DNA

Fetal cells

Biochemical analysis for

the presence of alleles

that cause many different

metabolic disorders

Growth forseveral days

in culture

medium

Karyotype analysis

(a)


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Chorionic Villus Sampling (CVS)

A method of sampling fetal chorionic cells byinserting a catheter through the vagina or abdominal

wall into the uterus

Used in diagnosing biochemical and cytogeneticdefects in the embryo

Usually performed in the eighth or ninth week ofpregnancy


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Chorionic Villus Sampling


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Chorionic

villi

Developing

placentaUltrasound to monitor

procedure

Developing

fetus

Bladder

Uterus

Chorion CatheterAmniotic

cavity

Rectum

(a)


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Variations in Chromosome Number

Changes in chromosome number or chromosomestructure can cause genetic disorders

Two major types of chromosomal changes can bedetected in a karyotypeA change in chromosomal numberA change in chromosomal arrangement


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Changes in Chromosome Number

PolyploidyA chromosomal number that is a multiple (3n or 4n)

of the normal haploid chromosomal number

AneuploidyA chromosomal number that is not an exact multiple

of the haploid number


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Polyploidy Changes the

Number of Chromosome Sets

TriploidyA chromosomal number that is three times the

haploid number, having three copies of all autosomes

and three sex chromosomes

TetraploidyA chromosomal number that is four times the haploid

number, having four copies of all autosomes and four

sex chromosomes


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A Triploid Karyotype


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Keep In Mind

Polyploidy results when there are more than twocomplete sets of chromosomes

A l id I l th G i L f


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Aneuploidy Involves the Gain or Loss of

Individual Chromosomes

MonosomyA condition in which one member of a chromosomal

pair is missing; one less than the diploid number (2n

1)

TrisomyA condition in which one chromosome is present in

three copies, and all others are diploid; one more

than the diploid number (2n + 1)


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Causes of Aneuploidy

Nondisjunction The failure of homologous chromosomes to separateproperly during meiosis

N di j ti i M i i I L d t


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Nondisjunction in Meiosis I Leads to

Aneuploidy


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Extra

chromosome(n + 1)Nondisjunction

Extra

chromosome(n + 1)

Missing

chromosome(n 1)

Missing

chromosome(n 1)

Meiosis I Meiosis II Gametes

(a)


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Nondisjunction

Extra

chromosome(n + 1)Normal division

Missing

chromosome(n 1)

Normal

(n)

Normal(n)

Meiosis I Meiosis II Gametes

(b)


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Effects of Monosomy and Trisomy

Autosomal monosomy is a lethal condition Eliminated early in development (spontaneous

abortion)

Some autosomal trisomies are relatively common Most result in spontaneous abortion Three types can result in live births (13, 18, 21)


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Trisomies in Spontaneous Abortions


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7.5Survey of 4,088

spontaneous abortions

5

4

3

2

Percentageoftrisomies

1

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22

Chromosome number


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Trisomy 13: Patau Syndrome (47,+13)

A lethal condition 1 in 10,000 births, mostdie within 1st month

Usually have polydactyly, eye

defects, severe brain, nervoussystem & heart defects


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Trisomy 18: Edwards Syndrome (47,+18)

A lethal conditionSurvival only 2-4 mths

1 in 11,000 births 80% are females Very slow growth,Mental retardation,

Heart malformations


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Trisomy 21: Down Syndrome (47, +21)

Occurs in 1/800 births Trisomy 21 is the only

autosomal trisomy

that allows survival

into adulthood

Mental retardation

Characterized byepicanthic fold

(corner of eye)

large furrowedtongues

40% have congenitalheart defects

High risk of leukemia &Alzheimers disease

Few reach the age of 50


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Trisomy 21: Down Syndrome (47,+21)

Monosomy and trisomy involve the

loss and gain of a single chromosometo a diploid genome


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ANIMATION: Nondisjunction

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What Are the Risks


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What Are the Risks

for Autosomal Trisomy?

The causes of autosomal trisomy are unknown Factors that have been proposed include:

Genetic predisposition Exposure to radiation Viral infectionAbnormal hormone levels

Maternal age is the leading risk factor for trisomy 94% of nondisjunctions occur in the mother


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18

16

Risk forDown syndrome

14

12

10

8

Trisomy21/1,0

00births

6

4

2

15 20 25 30 35 40 >44

Maternal age(a)

35


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35

30

Maternal age andtrisomic conceptions

25

15

10

5

Percentageofclinicallyrecognizedp

regnancies

15 16 18 20 22 24 26 28 30 32 34 36 38 40 42

(b)Maternal age


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Why is Maternal Age a Risk Factor?

Meiosis is not completed until ovulation Intracellular events may increase risk of

nondisjunction, resulting in aneuploidy

Maternal selection Embryo-uterine interactions that normally abort

abnormal embryos become less effective

Age of the mother is the best known risk factor fortrisomy


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Aneuploidy of the Sex Chromosomes

More common than autosomal aneuploidy Can involve both X and Y chromosomesA balance is needed for normal development

At least one copy of the X chromosome is requiredfor development Increasing numbers of X or Y chromosomes causes

progressively greater disturbances in phenotype and

behavior


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Turner Syndrome (45,X)

Monosomy of the X chromosome that results infemale sterility. Other phenotypic characteristics butotherwise normal.

Fig 6.20


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Klinefelter Syndrome (47, XXY)

Individuals (males) have some fertility problems butfew additional symptoms

Fig 6.22


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XYY Syndrome (47,XYY)

Affected individuals are usually taller than normaland some, but not all, have personality disorders

Changes in thenumber of sex

chromosomes haveless impact than

changes in

autosomes

Structural Alterations


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Structural Alterations

Within Chromosomes

Changes in the structure of chromosomes Deletion

Duplication Translocation Inversion


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Structural Changes in Chromosomes


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Deletions

Deletions involve loss of chromosomal material Deletions of chromosomal segments are associated

with several genetic disorders

Cri du chat syndrome Prader-Willi syndrome

Deletion in Chromosome 5 and cri du chat


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Deletion in Chromosome 5 and cri du chat

syndrome

Associated with an arrayof malformations, themost characteristic ofwhich is an infant cry that

resembles a meowing cat

due to defects in the

larynx

By comparing the regiondeleted with its associated

phenotype, investigators

have identified regions of

the chromosome thatcarry genes involved in

developing the larynx.

T l i


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Translocations

Translocation involves exchange of chromosomeparts

Often produces no overt phenotypic effects Can result in genetically imbalanced and aneuploid

gametes

Chromosomes can lose, gain, or rearrangesegments

R b t i T l ti


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Robertsonian Translocation

A translocation resulting in Down syndrome Robertsonian translocation makes Down syndrome a

heritable genetic disease

Potentially present in one in three offspring

Robertsonian Translocation and Down


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Robertsonian Translocation and Down

Syndrome

ANIMATION I i


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ANIMATION: Inversion



ANIMATION T l ti


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ANIMATION: Translocation



ANIMATION D li ti


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ANIMATION: Duplication



What Are Some


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Consequences of Aneuploidy?

Aneuploidy is the leading cause of reproductivefailure in humans

Results in miscarriages and birth defectsAneuploidy also is associated with many cancers

Chromosomal Abnormalities


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in Miscarriages

Other Forms of


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Chromosome Changes

Uniparental disomyA condition in which both copies of a chromosome

are inherited from a single parent

Copy number variationA situation in which a particular gene or chromosomal

region is present in multiple copies

Fragile sitesAppear as gaps or breaks in chromosome-specific

locations

Gene Imprinting, ie: Uniparental Disomy


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p g, p y

(UPD)

UPD is associated with several genetic diseases,also called gene imprinting

X-linked disordersAutosomal recessive disorders (Prader-Willi

syndrome, Angelman syndrome)

Prader-Willi syndrome: missing part of paternalchromosome 15, obesity, reduced mental ability &

muscle tone, little sex hormone production

Angelman syndrome: missing part of maternalchromosome 15, jerky movements, laughter (happypuppet syndrome)

Fragile Sites


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Fragile Sites

Appear as gaps or breaks in chromosomes One fragile site on the X chromosome is associated

with a common form of mental retardation in males

know as Fragile X Syndrome

Fragile Sites on the X Chromosome


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Fragile Sites on the X Chromosome

FRAX B


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FRAX BFRAX C

FRAX F

FRAX AFRAX D

FRAX E

The fragile sites

on the human X

chromosome.Sites B, C, and D

are common

sites and are

found on almost

all copies of the

X chromosome.A, E, and F are

rare sites;

expression of A

is associated

with fragile-X

syndrome.

Some fragilesites are

associated

with mental

retardation

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Cytogenetics Lesson 3