Dr. William Gartland 
May 11, 1979 
Page Two 
prevalent among E. col i strains associated with intestinal and extraintestinal 
disease, respectively, and all seem to be derepressed such that all cells that 
possess them have the F pi 1 i necessary for M13 infection. Fourth, while in- 
fection of these M13-sensitive hosts leads to persistent and continual release 
of M13 without death, infection of x-infectable hosts usually leads to death 
and lysis because of mutations present in most X vectors used in EK1 host-vector 
systems. 
In addition to the four factors enumerated above, there is an additional factor 
that may further reduce the biological containment of cloning systems using M13 
vectors and hosts with derepressed conjugative plasmids. A number of people 
have previously tried to determine whether M13 or fl DNA is transferred to an F~ 
strain as a consequence of conjugation promoted by the conjugative plasmid present 
in the M13- or fl-infected donor. Results were usually equivocal but recombinant 
technology should permit definitive conclusions to be made as to whether genetic 
information contained on an M13 or fl vector might be conjugational ly transmitted 
to another microorganism since one could easily measure conjugational transfer 
of a recognizable, selectable inserted gene in the M13 or fl vector. In the 
absence of data to disprove this route of transmission, I can only infer that it 
probably occurs and would contribute to an unreasonably high probability for 
dissemination of any recombinant DNA cloned on such phage vectors to other micro- 
organisms should the infected bacterial host or the vectors escape the laboratory 
environment. 
The proposed redefinition of EK1 also creates a loophole. For example, the pro- 
posed redefinition would allow cloning with a X vector capable of lysogenization 
into a host containing an F' gal' *' attx + over a A[ ga1 - attx] deletion mutation. 
This would permit easy transfer of the F' containing the recombinant X vector to 
a diversity of K- 12 hosts, a useful feature for screening functions of the re- 
combinant inserts, but would also permit efficient dissemination of the F* plasmid 
containing foreign genes to a diversity of microorganisms in nature. Although 
one could argue that zygotic induction would decrease the frequency of such dis- 
semination, zygotic induction is seldom complete and there are little or no data 
to my knowledge that indicate that this phenomenon occurs when X prophage is con- 
jugationally transferred to wild-type enteric isolates. On the contrary, L. S. 
Baron's group has found that zygotic induction is not detectable in most inter- 
generic matings with E. coli K-12 donors. 
Undoubtedly, others can think of additional ways to use the loophole created by 
the proposed redefinition of EK1. This fact and the poor biological containment 
properties of the M13 and fl host-vector systems, are the basis for my opposition 
to the proposed redefinition. In addition, the proposed revision can be ob- 
jected to based on historical and philosophical considerations. As you recall, 
after many attempts by the Recombinant Advisory Committee to distinguish between 
novel and non-novel in the definition of recombinant DNA, this approach was 
eventually discarded in favor of keeping the definition "pure" and to include 
in the Guidelines a category of exemptions to exclude non-novel recombinants from 
further consideration. Other provisions were also included in the Guidelines to 
permit modifications stemming from new information and situations. I thus 
[ 261 ] 
