-5- 
deficient background, etc., for example, £. col i . This experiment would be 
subject to regulation. Therefore, in attempting to introduce the recombinant 
molecules from the "colony bank" B into the various mutants of the source 
organism A., the investigator is de facto (and absolutely ) restricted to 
transformation since the current guidelines, as well as the proposed revisions, 
effectively prohibit conjugative mobilization by not allowing for the simul- 
taneous presence of conjugative and "novel" recombinant plasmids in the same 
host. The paradox is obvious: it is apparently all right to clone DNA from 
organism A using A itself, or another organism with which A exchanges chrom- 
osomal DNA, as the host, even though these may contain conjugative plasmids 
(free or integrated), or using a conjugative plasmid as a vector, and to trans- 
fer or mobilize such recombinant molecul es fro m these hosts to organisms X, 
Y, or Z with which A exchanges chromosomal DNA , since no "novel" recombinant 
molecules (as defined in the proposed revision) are involved in any of the 
steps mentioned; it is not all right, however, to perform the conjugative 
mobilization of the same molecules from another cloning host B, which does 
not exchange chromosomal DNA with Asunder any circumstances ! 
The P + HV containment levels assigned under the Guidelines are presumably 
commensurate with the probability that host B may be converted into a path- 
ogen. Therefore, it is the secondary biohazards of option (i) and (ii) 
above that must be compared. I know of no general rules that suggest that 
the intrinsic escape potential of recombinant plasmids from various unmodified 
(HV-1) procaryotic (Gram negative) primary cloning hosts, such as organisms A 
and B above, is not likely to be comparable. Granted, the likelihood of sec- 
ondary transmissions from such systems will be habitat-dependent. But in the 
absence of experimental data, habitat-dependent secondary transmission from 
[Appendix A — 114] 
