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Human

Chromosomes
and
Karyotyping
Karyotype

The entire chromosome complement of an
individual organism or cell as seen during
mitotic metaphase.


K562 Mammalian Cell Line
Myelogenous Leukemia Cell Line

Contains the Philadelphia Chromosome
A Translocation between the long arms of Chromosome
9 and Chromosome 22

Excellent in vitro growth characteristics
Translocation
The Relocation of a Chromosomal Segment to a
Different Position in the Genome.
Genetic Exchange of Chromosomal Material from
one Chromosome to Another.
Reciprocal Translocation – Equal Exchange of
Genetic Material between Two Chromosomes
Non-reciprocal Translocation – Unequal Exchange
of Genetic Material between 2 Chromosomes.
The Philadelphia Chromosome
A Kayrotype Containing the
Philadelphia Chromosome
Fluorescent In-Situ Hybridization (FISH)
identification of Human Chromosomes -
"Chromosome Painting"
Fluorescent In-Situ Hybridization (FISH)
Identification of Human Chromosomes -
"Chromosome Painting"
DNA Fragments with a Specific Sequence
are Labeled with a Fluorescent Dye
Labeled Fragments are allowed Hybridize to
Chromosomes therefore Binding to
Specific Complementary Sequences
Specific Sequences of DNA in the Genome
are Identified by Fluorescence.
Genetic Anomalies
Karyotyping allows the identification of
Genetic Anomalies
Simple Counting of Chromosomes can show
Genetic Disorders
Down’s Syndrome or Trisomy 21 easily
observed by the presence of 3 Chromosomes
21
Klinefelter’s Syndrome (XXY) will reveal 3 Sex
Chromosomes, 2 X’s and 1 Y
Genetic Anomalies
Euploidy – a cell having any number of
complete chromosome sets. In Humans
the complete set has 46 Chromosomes.
Aneuploidy – A cell having a chromosomal
number that differs from the normal
number of chromosomes for that species
by a small number of chromosomes.
Aneuploid Situations
Nullisomy – loss of one homologous pair (2N – 2)
Monosomy – loss of a single chromosome (2N – 1)
Turner’s Syndrome X0
Trisomy – a single extra chromosome (2N + 1)
Trisomy 21 – Down’s Syndrome
Klinefelter’s Syndrome - XXY
Tetrasomy – two extra of a single chromosome
(2N + 2)

Genetic Anomalies
Deletion of Genetic Material

Duplication of Genetic Material

Translocation of Genetic Material
Reciprocal Translocation
Non-reciprocal Translocation
Ploidy Levels
Diploidy – An organism having 2N number
of Chromosomes.
Monoploidy – An organism having 1N
number of Chromosomes. Useful in plant
Breeding.
Triploidy – An organism having 3N number
of Chromosomes.
Polyploidy – An organism with more than 2N
number of Chromosomes
Cell Preparation
Colchicine Treatment
Treat cells several hours before preparation

Colchicine interrupts spindle fiber formation
therefore no spindle fiber apparatus can form

Arrests cell cycle in Metaphase (Mitosis)

Cell Preparation
Incubate in 0.75M KCl
Hypotonic Solution
Swells Cells to almost Bursting

Incubate in Carnoy’s Fixative
Methanol:Glacial Acetic Acid 3:1
Fixes Cells by Cross-linking Proteins

Cell Preparation
Stain with Giemsa Stain
DNA Specific Stain
Heterochromatin, condensed inactive
DNA, stains more darkly.
Euchromatin, relatively uncoiled DNA, is
active and stains lightly.
Mitosis

Banding Patterns
Chromosomal Proteins are Partially
Digested using the Protease Trypsin.
Partial Digest reveals Bands along the Arms
of the Chromosome which have a Specific
Pattern.
There is an International Agreement as to
the Human Banding Pattern.

Banding Patterns in Human Chromosomes

Identifying Chromosomes
Chromosomes are arranged by size and
centromere location.
Chromosomes are categorized into 7 size
groups, A – G.
Chromosomes are further categorized by
Centromere location.
Centromere Locations
Metacentric – centromere in center of
chromosome arms.
Submetacentric – centromere is located just
off center of chromosome arms.
Acrocentric – centromeres located even
further from center of chromosome arms.
Telocentric – centromeres located near the
end of the chromosome arms.
Clinical Uses of Karyotyping
• Identification of Genetic anomolies
• Prenatal Screening of fetus
– Detect genetic anomolies before birth such as
Down’s Syndrome (Trisomy 21).
– This becomes more important as the age of
the mother increases. Above age 36 years is
the age where testing is recommended.
Here’s what you’ll see.