1. Chromosomal Structure and Chromosomal Mutations
Chapter 8
Chromosomal Structure and Chromosomal Mutations

2. 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.

3. 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

4. Types of Mutations
Genomic: abnormal chromosome number (monosomy, polysomy, aneuploidy)
Chromosomal: abnormal chromosome structure
Gene: DNA sequence changes in specific genes

5. 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

6. Chromosome Morphology
Telomere
Short
arm (p)
Arm
Centromere
Long
arm (q)
Telomere
Metacentric Submetacentric Acrocentric

7. Defining Chromosomal Locationp q
Arm
Region
Band
Subband 2 1 3 4 5
2, 3 17 .
Chromosome 17

8. 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

9. Cell Division Cycle Interphase (11–30 nm fibers) MetaphaseG1 S G2 M
Metaphase
(300–700 nm fibers)
Mitosis:
Prophase
Anaphase
Metaphase
Telophase

10. 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

11. Visualizing Metaphase Chromosomes (Banding)
Giemsa-, reverse- or centromere-stained metaphase chromosomes
G-Bands R-Bands C-Bands

12. Karyotype
International System for Human Cytogenetic Nomenclature (ISCN)
46, XX – normal female
46, XY – normal male
G-banded chromosomes are identified by band pattern.

13. Normal Female Karyotype (46, XX) (G Banding)

14. Normal Female Karyotype (High-Resolution G Banding)

15. Chromosome Number Abnormality Aneuploidy (48, XXXX)

16. Chromosome Number Abnormality Trisomy 21 (47, XX, +21)

17. Chromosome Structure Abnormalities
Translocation
Deletion
Insertion
Inversion
Isochromosome
Ring
chromosome
Derivative

18. Chromosome Structure Abnormality: Balanced Translocation 45, XY, t(14q21q)

19. 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)

20. Fluorescent in situ Hybridization (FISH)
Metaphase FISH
Chromosome painting
Spectral karyotyping
Interphase FISH

21. 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).

22. 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

23. FISH Probes Telomere-specific probes (TEL)
Hybridize to subtelomeric regions
Detect subtelomeric deletions and rearrangements
Probe binding site
Telomere
100–200 kb –20 kb
Unique sequences
Telomere associated repeats
(TTAGGG)n

24. Genetic Abnormalities by Interphase FISH LSI Probe
Greater or less than two signals per nucleus is considered abnormal.
Cell
nucleus
Normal diploid signal
Trisomy or insertion
Monosomy or deletion

25. Structural Abnormality by Interphase FISH LSI Probe (Fusion Probe)

26. Structural Abnormality by Interphase FISH LSI Probe (Break Apart Probe)

27. Translocation by Metaphase FISH WCP Probe (Whole-Chromosome Painting)

28. 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.