chapter 23
Structure and Properties of DNA
FIGURE 23-15
G enetic fingerprint o f m onozygotic tw ins (colum ns 2 and 3) and their
parents (colum ns 1
and 4). The arrows indicate hypervariable sequences
inherited from the father. Sim ilar hypervariable sequences can be identified
from the mother. [Reproduced with perm ission from A. J. Jeffreys,
V. W ilson, and S. L. Thein: Individual-specific “fingerprints” o f hum an
N a tu re
316, 76 (1985). © 1985 by M acm illan M agazines Ltd.]
that the sequences can be compared in order to eliminate
One sequencing technique is the
Sanger method
veloped by
Fred Sanger)
which uses dideoxynucleotides
that stop chain elongation at the site of their incorporation.
The four dideoxynucleotide substrates are labeled with
radioactivity and used in four separate reactions
in vitro
which a single-stranded DNA template is copied. A series
of radioactive DNA fragments are separated according to
length by agarose gel electrophoresis (fragments differing
by only one nucleotide are separated), and the DNA se-
quence can be read directly from the pattern of radioactive
bands in the gel.
Another sequencing technique is the
(developed by Alan Maxam and Walter Gilbert).
In this method, a single DNA strand is labeled at the 5'
end with radioactive phosphorus (P32). The radioactive
DNA is divided into four portions and each one is exposed
to different chemical reactions. Each reaction causes a 5'
cleavage adjacent to either
1. AorT,
2. G alone,
3. C or T, or
4. C alone.
The reactions are carried out for a short time so that, on
average, only one cleavage occurs in each DNA molecule.
This produces a set of DNA fragments (one set for each
reaction), whose length identifies that position of a par-
ticular base. For example, a fragment containing 19 nu-
cleotides in the G-only reaction mixture identifies G at
position 20 from the 5' end. Similarly, a fragment con-
taining 27 nucleotides present in the C or T reaction but
not in the C-only reaction indicates that T is at position
28. The lengths of DNA fragments are determined by
polyacrylamide gel electrophoresis,
a technique that can
separate DNA fragments differing in length by only one
Gene Therapy
One expected advance in medical treatment is the use of
gene therapy to ameliorate or cure a variety of inherited
disorders as well as diseases caused by somatic mutations
such as cancer. The goal of gene therapy is the replace-
ment of a defective, disease-causing gene in an individual
by normal, functioning copies. In essence, gene therapy
is a novel form of drug therapy; it uses the biochemical
capacity of a patient’s cells to synthesize the therapeutic
agent from the introduced gene.
The first attempts at gene therapy were directed at try-
ing to correct
severe combined immunodeficiency (SCID)
which is caused, in some cases, by a deficiency of adeno-
sine deaminase (ADA), that is expressed in all tissues.
ADA deaminates both adenosine and deoxyadenosine
and, in the absence of ADA, deoxyadenosine accumulates
in cells. Deoxyadenosine can be phosphorylated by the
enzyme deoxcytidine kinase to produce deoxyadenosine
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