Biol 107, Spring 2005

Human Karyotype Exercise

Occasionally chromosomal material is lost or rearranged during the formation of gametes or
during cell division of the early embryo. Such changes, primarily the result of nondisjunction or
translocation, are so severe that the pregnancy ends in miscarriage – or fertilization does not
occur at all. Rarely, if the chromosomal change is relatively minor, the resulting embryo will
survive, but it may develop abnormally in some way. It is estimated that one in 156 live births
have some kind of chromosomal abnormality.

Some of the abnormalities associated with chromosome structure and number can be detected by
a test called a karyotype. A karyotype can show prospective parents whether they have certain
abnormalities that could be passed on to their offspring, or it may be used to learn the cause of a
child’s disability. Karyotypes can also reveal the gender of a fetus or test for certain defects
through examination of cells from uterine fluid – a procedure called amniocentesis – or through
sampling of placental membranes. Over 400,000 karyotype analyses are performed each year in
the U.S. and Canada.

To create a karyotype, chromosomes from a cell are stained and photographed. The photograph
is enlarged and cut up into individual chromosomes. The homologous pairs are identified and
arranged in order by size (with the exception of the sex chromosomes; these appear last). These
tests are typically done on a sample of blood, although any body cell could be used. The cell
must be undergoing mitosis – preferably in metaphase – so that the chromosomes are replicated,
condensed, and visible under a microscope.

"Spread" of human chromosomes A karyotype of

from a single cell human chromosomes
Some chromosomal abnormalities that can be detected by karyotyping are listed below:

Abnormality Effects

Extra chromosome 21 Associated with mental retardation, characteristic facial

Down Syndrome features and stature, heart defects, respiratory infection,
leukemia, and Alzheimers disease; occurs in 1 in 700 births
in U.S.

XXY Typically sterile males with abnormally small testes, some

Klinefelter Syndrome female characteristics, normal intelligence; occurs in 1 in
2000 births

XYY Affected individuals tend to be taller as a group,

Double Y Syndrome normal intelligence

XXX Distinguished from normal XX females only through

karyotyping; occurs in 1 in 1000 births

XO - missing sex chromosome Sterile females with immature sex organs, normal
Turner Syndrome intelligence; occurs in 1 in 5,000 births

Partial chromosome 5 Associated with small head, characteristic cry and facial
Cru de chat Syndrome features, death in infancy or early childhood

Abnormal X chromosome Common genetic cause of mental retardation

Fragile X Syndrome

Today in lab you will assemble a human karyotype. In this case, the chromosomes are from a
white blood cell (leucocyte) of a living adult. Note that the chromosomes in the photograph are
in their doubled state - two chromatids connected by a centromere. However, the two
chromatids are so close together that you cannot distinguish between them. An indentation often
appears where the centromere connects the two. This entire structure is one chromosome. (In
some other preparations the chromatids may be spread in an "x" shape.)

---chromatid

Chromosome--- = ---centromere
as it appears
in photograph chromatid ---
Remember that normal human cells contain 46 chromosomes – 23 contributed by the mother and
a corresponding set of 23 that came from the father. A karyotype matches up the 23 pairs. Three
elements are considered in matching a chromosome with its homologue: length, centromere
location, and the banding pattern produced by staining. (A common misconception is that each
dark band represents a single gene. One thin band may actually contain hundreds of genes.)

Homologous pairs
share these attributes: length —
—centromere location

—banding pattern

Procedure – Work in Pairs

NOTE: Abnormalities will be related to chromosome number. None of the subjects will have
more than one abnormality. Some subjects may be normal.

1. Your TA will give you a blank karyotype form and an enlarged photograph of a spread of
chromosomes from a human leucocyte. Record on your karyotype form the Subject Number
that appears at the top of your sheet of chromosomes.

2. Count the chromosomes on your sheet (this initial count should give you a preliminary idea
about the nature of your subject). Now carefully cut out each chromosome (leave white
space around each chromosome rather than trying to cut exactly on the margin of the image).
Share the work with your partner. Be careful not to lose any chromosomes.

3. Find the homologous pairs by matching length, the position of the centromere, and the
banding patterns if present. All three of these elements may be needed to make an accurate
match.

4. Arrange the pairs on the karyotype form in order from longest to shortest, with the exception
of the sex chromosomes, X and Y. These appear last. The sex chromosomes are obviously
not a homologous pair: the X chromosome is of medium length, intermediate between
chromosomes 4 and 5; the Y is one of the smaller chromosomes, similar in length to
chromosome 14.

5. Lightly tape the chromosomes in place. Don't use a lot of tape initially – have your TA
check your work, then tape more securely.
Analysis

Does your karyotype have an abnormal chromosome number? This may be because
chromosomes were lost or cut incorrectly. It is also possible that the karyotype is of a person
with a chromosomal abnormality. How can you decide which is the case? Hint: These
chromosomes were sampled from a living adult. Humans cannot survive to adulthood if they are
missing any autosomes. Some females may survive with just one X chromosome (XO).
Humans can survive with an extra X or Y chromosome (XXX, XXY, XYY). They may also live
to adulthood with an extra chromosome 21 (Down Syndrome).

Record the number of chromosomes present on your karyotype and the gender of the subject. If
there is an abnormal number of chromosomes (more or less than 46), note which ones are
missing or in excess. Your subject may have one of the abnormalities listed earlier. Which
abnormality do you think he or she has?

Before you leave, be sure to look at the demonstration slide of chromosome spreads from a
human leucocite and examine the karyotypes on display in lab. Some of the karyotypes exhibit
structural abnormalities in individual chromosomes. Cytogeneticists can detect even smaller
changes than these - down to slight differences in individual bands.

Questions to think about:

How might a cell come to have an abnormal number of chromosomes?

Why is it necessary in karyotyping to use cells that are undergoing cell division?

What are some limitations of the karyotype as a test for genetic abnormalities?