NOTICES 
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During the past 30 years the struc- 
ture of DNA molecules has been stud- 
ied intensively, and it can now be de- 
scribed in much detail. The molecule 
may be compared to a long-twisted 
stepladder with thousands to millions 
of rungs. A short piece of DNA is rep- 
resented in Figure 2. 
Figure 2 
The sides of the ladder are formed 
of sugar molecules (dexoyribose) at- 
tached end to end through phosphate 
groups. At right angles to each sugar 
molecule is one of four possible 
bases— adenine, guanine, thymine, and 
cytosine. The precise sequence of 
these bases, the rungs of the ladder, 
codes the information content. The 
"reading” of the code contained in the 
sequence of bases results in the forma- 
tion of proteins, which in turn carry 
out most of the essential functions of 
the ceil. 
A gene is a portion of the DNA mole- 
cule which codes for the manufacture 
of a protein. In higher organisms, 
much of the DNA may not serve as 
genes in this sense, but may regulate 
the activity of nearby genes. It is pos- 
sible to break open cells and isolate 
DNA, free of other cellular constitu- 
ents. 
The formation of "recombinant 
DNA” in the laboratory was made pos- 
sible by a series of discoveries. W. 
Arber and D. Dussoix, in 1962, showed 
that bacteria contain substances called 
restriction enzymes. Serving to defend 
the bacteria against viruses, these en- 
zymes can split foreign DNA molecules 
into specific fragments. R. Yoshimori, 
in H. Boyer’s laboratory, isolated a re- 
striction enzyme that was later found 
to split DNA into fragments whose 
ends stick together when they touch. 
In 1973 S. Cohen and others succeeded 
in combining genes of different species 
and introducing them into bacteria. 
Then they grew the bacteria in cul- 
tures, multiplying the combined char- 
acteristics. 
The capabilities sketched here— to 
split DNA selectively, to recombine it 
by virtue of "sticky” ends, to reintro- 
duce it into cells, and to cultivate the 
cells— constitute the recombinant 
DNA technique. 
In the recombinant DNA experi- 
ments that are the subject of the NIH 
guidelines, the DNA can be derived 
from widely divergent sources. DNA 
from one of the sources may serve as a 
carrier, or “vector,” for the insertion 
of the recombined DNA into a cell, or 
“host.” The vector may be a plasmid, 
usually derived from the same species 
as the host, or it may be a virus. The 
DNA to be inserted is called the "for- 
eign” DNA. When a large mixture of 
DNA fragments from the foreign 
source is used in the joining, the ex- 
periment is referred to as a "shotgun” 
experiment. In other instances, a par- 
ticular DNA fragment of interest will 
be purified and then incorporated in 
the vector. 
From a growth culture of the host 
cells, one containing the interesting 
DNA fragment is selected and allowed 
to multiply. The resulting population 
of identical cells is called a “clone.” In 
some experiments the DNA will be ex- 
tracted from the cells for study; in 
others, the properties of the cells 
themselves will be investigated. 
In the experiments discussed in the 
guidelines, the host cells are generally 
single-cell microorganisms such as bac- 
teria, or animal or plant cells that 
were originally obtained from living 
tissue but are grown as single cells 
under special laboratory conditions. 
The process of producing recombin- 
ant DNA molecules and introducing 
them into cells is illustrated in figure 
3. 
Figure 3 
I 
[Chromo-] ) 
QO O 
Host Cell 
The cell represented at the upper 
left contains chromosomal DNA and 
several separately replicating DNA 
molecules. The nonchromosomal DNA 
molecules can be isolated from the cell 
and manipulated to serve as vectors 
(carriers) for DNA from a foreign cell. 
Most DNA molecules used as vectors 
are circular. They can be cleaved, as 
shown, by enzymes (restriction endon- 
ucleases) to yield linear molecules 
with rejoinable ends. 
At the upper right is another cell, 
represented here as a rectangle. It 
serves as the source of the foreign 
DNA to be inserted into the vector. 
This DNA can also be cleaved by en- 
zymes. The rectangular cell could be 
FEDERAL REGISTER, VOL. 43, NO. 146— FRIDAY, JULY 28, 1978 
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