Chapter 8
Chromosomal Structure and Chromosomal
Mutations
Objectives
 Define mutations and polymorphisms.
 Distinguish the three types of DNA mutations:
genome, chromosomal, and gene.
 Diagram a human chromosome and label the
centromere, q arm, p arm, and telomere.
 Illustrate the different types of structural
mutations that occur in chromosomes.
 Show how karyotypes reveal chromosomal
abnormalities.
 Describe interphase and metaphase FISH
analyses.
Mutations and Polymorphisms
 Mutation: a permanent transmissable
change in the genetic material, usually in
a single gene
 Polymorphism: two or more genetically
determined, proportionally represented
phenotypes in the same population
Types of Mutations
 Genomic: abnormal chromosome number
(monosomy, polysomy, aneuploidy)
 Chromosomal: abnormal chromosome
structure
 Gene: DNA sequence changes in specific
genes
Chromosome Morphology
 Telomere: chromosome
ends
 Centromere: site of spindle
attachment
 Constriction of the
metaphase chromosome at
the centromere defines two
arms
 Nucleosome: DNA double
helix wrapped around
histone proteins
Chromosome Morphology
Telomere
Centromere
Arm
Short
arm (p)
Long
arm (q)
Telomere
Metacentric
Submetacentric
Acrocentric
Defining Chromosomal Location
Arm
Region Band Subband
2
p
1
1
2
1
1
1
2
1
2
q
3
2
Chromosome 17
3
2
1
2
1
5
4
3
2
1
4
3
1
2
3
1
2, 3
4
1
2
3
17q11.2
Chromosome Morphology Changes
During the Cell Division Cycle.
 DNA double helix: 2nm diameter
Interphase (G1, S, G2)
 Chromatin “beads on a string:” 11nm
 Chromatin in nucleosomes: 30nm
Metaphase (Mitosis)
 Extended metaphase chromosomes: 300 nm
 Condensed metaphase chromosomes: 700 nm
Cell Division Cycle
Interphase
(11–30 nm fibers)
G1
S
G2
M
Mitosis:
Prophase
Anaphase
Metaphase
Telophase
Metaphase
(300–700 nm fibers)
Visualizing Metaphase
Chromosomes
 Patient cells are incubated and divide in
tissue culture.
 Phytohemagglutinin (PHA): stimulates cell
division
 Colcemid: arrests cells in metaphase
 3:1 Methanol:Acetic Acid: fixes
metaphase chromosomes for staining
Visualizing Metaphase
Chromosomes (Banding)
 Giemsa-, reverse- or centromere-stained
metaphase chromosomes
G-Bands
R-Bands
C-Bands
Karyotype
 International System for Human
Cytogenetic Nomenclature (ISCN)
 46, XX – normal female
 46, XY – normal male
 G-banded chromosomes are identified by
band pattern.
Normal Female Karyotype (46, XX)
(G Banding)
Normal Female Karyotype
(High-Resolution G Banding)
Chromosome Number Abnormality
Aneuploidy (48, XXXX)
Chromosome Number Abnormality
Trisomy 21 (47, XX, +21)
Chromosome Structure
Abnormalities
Translocation
Deletion
Derivative
chromosome
Inversion
Insertion
Isochromosome
Ring
chromosome
Chromosome Structure Abnormality:
Balanced Translocation 45, XY, t(14q21q)
Fluorescent in situ Hybridization
(FISH)
 Hybridization of complementary gene- or
region-specific fluorescent probes to
chromosomes.
Interphase or metaphase
cells on slide (in situ)
Probe
Microscopic
signal (interphase)
Fluorescent in situ Hybridization
(FISH)
 Metaphase FISH
 Chromosome painting
 Spectral karyotyping
 Interphase FISH
Uses of Fluorescent in situ
Hybridization (FISH)
 Identification and characterization of
numerical and structural chromosome
abnormalities.
 Detection of microscopically invisible
deletions.
 Detection of sub-telomeric aberrations.
 Prenatal diagnosis of the common
aneuploidies (interphase FISH).
FISH Probes
 Chromosome-specific centromere probes (CEP)
 Hybridize to centromere region
 Detect aneuploidy in interphase and metaphase
 Chromosome painting probes (WCP)
 Hybridize to whole chromosomes or regions
 Characterize chromosomal structural changes in metaphase
cells
 Unique DNA sequence probes (LSI)
 Hybridize to unique DNA sequences
 Detect gene rearrangements, deletions, and amplifications
FISH Probes
 Telomere-specific probes (TEL)
 Hybridize to subtelomeric regions
 Detect subtelomeric deletions and rearrangements
Probe binding site
Unique sequences
Telomere
100–200 kb
3–20 kb
Telomere associated repeats
(TTAGGG)n
Genetic Abnormalities by
Interphase FISH LSI Probe
 Greater or less than two signals per
Cell
nucleus is considered abnormal.
nucleus
Normal diploid signal
Trisomy or insertion
Monosomy or deletion
Structural Abnormality by Interphase
FISH LSI Probe (Fusion Probe)
Structural Abnormality by Interphase
FISH LSI Probe (Break Apart Probe)
Translocation by Metaphase FISH
WCP Probe (Whole-Chromosome Painting)
Summary
 Mutations are heritable changes in DNA.
 Mutations include changes in chromosome number,
structure, and gene mutations.
 Chromosomes are analyzed by Giemsa staining and
karyotyping.
 Karyotyping detects changes in chromosome number
and large structural changes.
 Structural changes include translocation, duplication,
and deletion of chromosomal regions.
 More subtle chromosomal changes can be detected by
metaphase or interphase FISH.