Chapter 30

Cytogenetics

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chapter Overview 2

• Reasons for chromosome analysis

• Chromosome structure
• Chromosome identification
• Chromosome preparation
• Techniques of chromosome preparation and analysis
• Cytogenetic nomenclature
• Chromosome abnormalities
• Cancer cytogenetics
• Chromosomal microarray analysis
Reasons for Chromosome Analysis 3

• Karyotyping ordered to evaluate

• Mental retardation
• Infertility
• Ambiguous genitalia
• Short stature
• Fetal loss
• Risk of genetic or chromosomal disease
• Cancer
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Chromosome Structure 4

• Cell cycle
• Four stages
• G1 -the cell increases in mass in preparation for
cell division
• DNA synthesis - The period during which DNA
is synthesized
• G2 -The cell synthesizes proteins and continues
to increase in size
• Mitosis
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Chromosome Structure (Cont.) 5

• Chromosome architecture
• Double-stranded DNA
• Backbone
• Sugar
• Bases
• Purines
• Pyrimidines
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Continued
Chromosome Structure (Cont.) 6

• Chromosome architecture
• Chromatin
• Condensation
• Multiple levels of
coiling

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chromosome Identification 7

• Chromosome number in humans

• 2n = Diploid number
• Somatic cells—46 chromosomes
• n = Haploid number
• Gametes—23 chromosomes

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Continued
 Chromosome Size and Type 8

The three shapes of chromosomes are

metacentric, submetacentric, and acrocentric.
This figure also shows the position of the
centromere and telomere as well as the two
sister chromatids that comprise a metaphase
chromosome.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

 Techniques for Chromosome Preparation and 9

Analysis
• Chromosome preparation
• Cell characteristics
• High mitotic rate—bone marrow
• Stimulated to divide in culture—peripheral blood
• Harvesting procedures
• Mitogens added to stimulate cellular division (if needed)
• Cells arrested in metaphase
• Agent added to disrupt mitotic spindle fiber attachment
• Hypotonic solution added to spread chromosomes apart
• Fixation
• Dispersal on slides and aging by exposure to heat
Continued
Techniques for Chromosome Preparation and Analysis 10

(Cont.)

• Chromosome banding
• Greek meaning of chromo and soma
• Q banding
• G banding
• GTG banding
• C banding
• AG-NOR banding

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Techniques for Chromosome Preparation and Analysis 11

(Cont.)

• Metaphase analysis
• Chromosome number
• Banding pattern
• Karyotype
• Karyogram

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Techniques for Chromosome Preparation and Analysis 12

(Cont.)

• Fluorescence in situ hybridization

• Dividing and nondividing cells
• DNA or RNA probe labeled with a fluorophore
• Target DNA treated and washed
• Cells counterstained for visualization

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

 Fluorescence in Situ Hybridization
13

General protocol for fluorescence

in situ hybridization. PCR,
Polymerase chain reaction.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Cytogenetic Nomenclature 14

• Paris conference of 1971

• Short arms (p)
• Long arms (q)
• Numbering of bands
• Numbering of chromosomes
• 1 to 22 somatic
chromosomes
• X and Y sex chromosomes
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Chromosome Abnormalities 15

• Numeric abnormalities
• Aneuploidy
• Definition – abnormal number of chromosomes,
not a multiple of 23
• Examples: trisomy, monosomy
• Polyploidy
• Definition – Chromosomes number is greater
than 46
• Examples: triploidy has 69 chromosomes,
tetraploidy has 92
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Continued
Chromosome Abnormalities (Cont.) 16

 Triploid karyotype, 69,XXY.

Hypodiploid karyotype with 36

chromosomes (arrows indicate
missing chromosomes).
Chromosome Abnormalities (Cont.) 17

• Structural abnormalities
• Balanced versus unbalanced
• Structural rearrangements
• Inversions
• Deletions
• Isochromosomes
• Ring formations
• Insertions
• Translocations
• Duplications
Chromosome Abnormalities (Cont.) 18

 A, Pericentric inversion involves the centromere. B,

Paracentric inversion occurs in either the short or long
arm of the chromosome.
Chromosome Abnormalities (Cont.) 19

A, Interstitial deletion.
B, Isochromosome.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chromosome Abnormalities (Cont.) 20

C, Ring chromosome.
D, Insertion.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Cancer Cytogenetics 21

• Clinical applications
• Area of study focusing on nonrandom chromosome
abnormalities in many types of cancers.
• Suitable specimens
• Bone marrow is commonly used
• Structural rearrangements
• Defects are numeric abnormalities and structural
rearrangements on the chromosome
Continued
Cancer Cytogenetics 22

• Clone
• Is a cell population derived from a single progenitor.
• A clone exists if 2 or more cells contain same structural
abnormality
• Types of aberrations -
• Primary – cytogenic abnormality associated with a cancer
• Secondary – include other abnormalities

Continued
Cancer Cytogenetics (Cont.) 23

• Leukemia
• Characteristics
• Lymphoid or myeloid
• Main Categories
• Acute lymphoblastic leukemia (ALL)
• Acute myeloid leukemia (AML)
• Chronic lymphocytic leukemia (CLL)
• Chronic myelogenous leukemia (CML)
• Classification
• World Health Organization (WHO)
• French-American-British (FAB)
Continued
Cancer Cytogenetics (Cont.) 24

• Chronic myelogenous leukemia

• First malignancy to be associated with chromosomal abnormality
• 95% of patients have Philadelphia chromosome
• BCR, a gene on chromosome 22 joins ABL1, a gene on
chromosome 9. This causes cells to grow and become cancerous.
• Detection by FISH technology
• Monitoring is performed using peripheral blood

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Continued
Cancer Cytogenetics- CML
25

Normal bone marrow interphase Abnormal bone marrow interphase

cell hybridized with the BCR cell with one BCR (green) and one
(green) and ABL1 (red) genes ABL1 (red) signal
Cancer Cytogenetics (Cont.) 26

• Acute leukemia
• Recurrent
genetic
abnormalities
• ALL examples
• AML examples

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chromosomal Microarray (CMA) Testing 27

• Fluorescence-based molecular technique

• Submicroscopic analysis of genomic DNA
• High resolution
• Detection of chromosomal abnormalities
• Inherited disorders
• Emerging applications for cancer

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chromosomal Microarray (Cont.) 28

Chromosomal microarray diagram

demonstrating approximately a 32.4-
Mb gain of genetic material between
bands 7q21 and 7q31.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chromosomal Microarray (Cont.) 29

Chromosomal microarray diagram

demonstrating approximately a 4.6 Mb loss at
band 7q31.1.

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.

Chapter Review 30

• State some reasons why karyotyping is ordered?

• Draw and label the DNA molecule.
• What is the diploid number of chromosomes?
• Describe how a karyogram is determined?
• How does FISH detect abnormalities in chromosome
number and structure?
• How does analysis of chromosomes impact diagnosis,
prognosis, and therapy of leukemias?
Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.
Reticulocytes 31

Copyright © 2016, 2015, 2012 by Saunders, an imprint of Elsevier Inc.