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Federal Register / Vol. 46, No. 233 / Friday. December 4. 1981 / Notices 
general arguments about the ability of 
the-organism carrying recombinant DNA 
to compete successfully with 
nonrecombinant DNA organisms. 
There is a great deal of anecdotal 
information about the difficulties that 
people have maintaining recombinant 
DNA in bacterial strains; selective 
pressures must be applied continuously 
to maintain many plasmids. Little of this 
type of information has been published. 
One such experiment, reported by 
Cameron and Davis (1977). examines the 
fate of random £. coli and 
Saccharomyces cerevisiae fragments 
cloned into a bacterial virus vector and 
propagated for many cycles in E. coli. 
After about 25 cycles of growth, both 
sets of DNAs were reduced from the 
original diverse population to one or two 
dominant types which presumably have 
a growth advantage over all the other 
types. The authors also state that none 
of the yeast clones outgrow the parent 
vector (containing no recombinant DNA 
information). Therefore, most 
recombinants grown under these 
conditions will be quickly lost after the 
host organism begins to multiply. 
Evolutionary arguments suggest that 
an organism containing recombinant 
DNA information will be at a relative 
disadvantage; this is particularly true for 
complex organisms such as mammals. 
! Ayala (1977) points out that, for 
developmentally advanced organisms, 
new information must be coadapted to 
: the rest of the gene pool of that 
I organism; this is almost impossible to do 
1 with new information unless the new 
information is simply a different form of 
an already present gene. If. however, the 
new information is very similar to that 
which is already present, such a variant 
might have arisen by natural means and 
I the recombinant DNA containing 
j organisms will not be unique. Chances 
I for altering the evolution of simpler 
I organisms, such as prokaryotes, may be 
I somewhat greater, althou^ short 
generation times and relatively 
I economical use of DNA sequence 
information suggests that non-useful 
information will be rapidly lost. 
! Overproduction of one or a few products 
, would be expected to unbalance the 
I cell's metabolism. 
I b. Transmission into other potential 
hosts. There are three major 
, mechanisms for transfer of recombinant 
[ DSA from the original host to other 
I hosts encountered in the environment. 
' (1) For plasmids, in particular, the 
j conjugational mode (by mating. 
< involving cell-to-cell contact) may be 
I primary. Either conjugative transfer of a 
I self-transferring plasmid or mobilization 
I of non-conjugative plasmids may occur 
(Low Porter, 1978). At least for one 
class of vectors frequently used for 
recombinant DNA, those based on the E. 
coli plasmid pBR322, it has been 
demonstrated in vitro and in vivo that 
transfer by mobilization is greatly 
reduced (Dougan, G., Crosa, J.H., Falkow 
S., 1978; Levine. M.M., Kaper, J.B., et al., 
unpublished data) apparently because a 
segment necessary for mobilization was 
deleted in construction of the vector 
(Clark & Warren, 1979). Clearly the 
range of possible recipients will depend 
on the host range of the transferring 
plasmids. 
(2) Viral vectors, if they are intact, 
may readily transfer the recombinant 
DNA they carry to other sensitive hosts. 
If the host is killed in the process, the 
association will be only a temporary 
one. If the vector is defective (lacking an 
essential function), the function must be 
supplied by a helper virus for each new 
round of growth and infection. 
Therefore, if an appropriate helper is not 
commonly found in the environment, it 
is unlikely that the defective viral vector 
and its recombinant DNA will be 
disseminated. 
(3) Finally, cells are capable of taking 
up naked foreign DNA. For bacterial 
systems carrying out transformation, 
restriction systems will frequently 
degrade the incoming DNA if it is from a 
foreign strain, thereby reducing greatly 
the probabilities for stable incorporation 
of incoming recombinant DNA. In 
addition, it has been found that 
plasmids serve as a particularly poor 
source of DNA for transformation into 
some strains [B. subtilis] (Canosi et al., 
1978; Contente & Dubnau, 1979). Since 
DNA which is nonhomologous and is 
not competent to replicate by itself will 
not be recombined into the host 
chromosome or maintained in the 
cytoplasm, it may be reasonable to 
assume that naked recombinant DNA 
taken up by prokaryotes by 
transformation will not generate stable 
recombinant DNA containing organisms. 
In addition, at least in some 
environments, nucleases, pH and other 
environmental factors should interfere 
with the stability of naked recombinant 
DNA. In one experiment, bacterial DNA 
exposed to the diluted contents of rat 
intestine was rapidly degraded (Maturin 
and Curtiss. 1977). In other cases, 
however, in vivo DNA transformation 
has been observed, suggesting that not 
all DNA released into the animal tissues 
will be immediately destroyed 
(Ottolenghi-Nightingale, 1969). 
Take-up of DNA by animal cells in 
tissue culture is not an efficient process 
under optimized laboratory conditions 
(Scangos & Ruddle, 1981). Moreover, for 
an effective cycle which will lead to 
wide-scale dissemination, the DNA 
would have to be stably integrated into 
the germ line of an intact organism. 
Alternatively, some constant source 
pool, such as infecting bacteria or viral 
recombinants may continue to provide a 
source of the DNA. 
Individual plant cells, since they can 
be regenerated into complete plants, 
could disseminate DNA if it were 
integrated into nuclear or organelle 
DNA. However, this may also require a 
constant bacterial or viral source pool in 
plant cells. 
3. Harm. 
Let us assume for the moment that the 
organisms we have created via 
recombinant DNA are in fact unique, 
and have had the opportunity and 
ability to establish themselves in the 
environment. Will these organisms have 
the ability to cause harm, either to 
ourselves, to other animals, or to plants, 
microorganisms and the environment? 
We will consider here some general 
arguments about the roles recombinant 
DNA can play, and some of the specific 
cases which people have considered to 
be areas of concern. 
a. Breaching prokaryotic-eukaryotic 
barriers: evolutionary considerations. 
Prominent among the concerns 
expressed during the first few years of 
recombinant DNA technology was the 
fear of “breaching the barriers" to 
permit recombination between distantly 
related organisms. This concern, 
articulated most prominently by 
Sinsheimer (1975, 1976 a, b) and by 
Chargaff (1976), implies that the fertility 
barriers that evolved as organisms 
diverged during evolution arose to 
prevent the creation of new. dangerous 
species. Davis argues that this view 
"turns evolutionary principles upside 
down. Evolution has indeed established 
fertility barriers between species. But 
these barriers do not function to prevent 
the formation of monsters that might 
take over in the Darwinian struggle; 
They prevent wasteful matings that 
would produce only unbalanced 
monstrosities, unable to survive” (Davis, 
1976, 1977). 
In addition, information based 
primarily on the recombinant DNA 
analysis of eukaryotic genomes makes it 
clear that the mechanisms for 
processing information from DNA into 
protein differ greatly between many 
eukaryotes and prokaryotes (Breathnach 
& Chambon. 1981; Revel & Groner, 1978). 
Therefore, it is difficult to imagine that 
for most cases the “breaching of the 
brarrier" will result in much more than 
adding silent DNA sequences into the 
host cell. In the absence of any 
expressed function, it is difficult to 
hypothesize either a beneficial or 
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