by molecular diagnostic studies. When a disease is
always caused by the same mutation, this is a very
accurate way to ascertain carrier status. Sickle cell
anemia is an example.
When more than one mutation can cause a
genetic disease, carrier detection by molecular
methods is usually somewhat less sensitive because it
is usually only practical to detect the most prevalent
mutations. In the case of Tay-Sachs disease, for
instance, the three mutations that cause 98 percent of
the disease among Ashkenazi Jews are detected in
routine testing. That means that the sensitivity of
the study is, at best, 0.98 in this subpopulation. The
sensitivity is lower in the general population where
there is a greater variety in the mutations that cause
the disease.
There are a large number of mutations that cause
hemophilia and no one mutation is responsible for
more than a small fraction of the cases. As a conse-
quence, molecular diagnostic evaluation of carrier
status in hemophilia is very involved and is usually
only performed at larger medical centers. If there
are a number of family members with the disease,
gene linkage studies can be conducted with the hope
of identifying a DNA sequence polymorphism that is
linked reliably to the disease-producing mutation. If
family studies are unrewarding, or if there are not
enough evaluable family members with the disease,
the DNA of the suspected carrier can be screened for
abnormalities. Deletions and nucleotide substitu-
tions that disrupt the site of a restriction enzyme can
be identified by Southern blot analysis (Peake 1995).
Deletions are demonstrated by probing restriction
enzyme-treated DNA with the complete factor VIII
cDNA. Nucleotide substitutions are found by analy-
sis of the fragment patterns of the exons, intron/exon
borders, and 5’ and 3’ non-coding regions of the
factor VIII gene following incubation with each of
many different restriction enzymes. Mutations that
do not disrupt restriction enzyme sites can be
screened for using mismatch detection techniques
(Ferrari
et al.
1996, Nollau
et al.
1997). Such
studies do not reveal a defect in approximately half
of the cases in which the mutation results in no gene
product. Most of these cases are caused by an inver-
sion of the distal portion of the X chromosome that
leads to disruption of the factor VIII gene. The
location of the inversion is always the same so it can
be easily detected. Using a probe for the DNA locus
responsible for the inversion, Southern blot analysis
of the DNA fragments generated by the restriction
enzyme
Bcl
I reveals fragment polymorphisms that
are characteristic for the inversion (Peake 1995).
Prenatal detection of genetic disease
Prenatal laboratory screening for genetic disease
is limited to studies that can be carried out on speci-
mens that can be collected with very low risk to the
mother and fetus. By far the safest specimen to
obtain is maternal blood.
In cases in which the fetus has an increased
probability of having a genetic disease, the physician
and parents may agree to pursue more definitive
laboratory studies. As most genetic diseases can
only be diagnosed with high reliability using labora-
tory studies performed on cells, this amounts to a
decision to obtain and study fetal cells. This neces-
sitates invasive specimen collection techniques that
entail some risk to the mother or fetus.
Maternal blood.
The placental trophoblasts
synthesize various hormones and nonhormonal
proteins. All of these substances are able to diffuse
into the maternal circulation (Chard 1991). The
concentrations of the placental products in the mater-
nal plasma reflect the rate of synthesis of the respec-
tive products in the placenta. Thus, if a genetic
disease alters the rate of placental synthesis of a
product, the concentration of the product will be
altered in the maternal plasma. In this way, placen-
tal products can be used as diagnostic markers for
some genetic diseases. In trisomy 21, for example,
the placental production of intact human chorionic
gonadotrophin and its free beta subunit are increased
(Newby
et al.
1997) resulting in increased concen-
trations of these products in the maternal plasma.
The placenta also permits the ready exchange of
lipophilic substances between the fetal circulation
and the maternal circulation (Chard 1991). Thus,
any lipophilic substance that accumulates in the fetal
plasma as a result of a genetic disorder will also
distribute into the maternal plasma and may reach
concentrations that allow it to be used as a diagnostic
marker. In addition, certain substances are trans-
ported from the fetal plasma into the maternal
plasma by the placenta. A transported substance can
serve as a diagnostic marker if the rate of transport
of the substance is altered by a genetic disease. The
placental transport of
α
-fetoprotein appears to be
diminished in trisomy 21 (Newby
et al.
1997)
leading to decreased
α
-fetoprotein concentrations in
the maternal plasma. There is substantial overlap of
the marker reference frequency distributions in
Genetic Disease
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