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III.  Experimental Guidelines

A general rule that, though obvious, deserves statement is that the

level of containment required for any experiment on DNA reco'mbinants shall

never be less than that required for the most hazardous component used to

construct and clone the recombinant DNA (i.e., vector, host, and inserted

DNA).  In most cases the level of containment will be greater, particularly

when the recombinant DNA is formed from species that ordinarily do not ex-

change genetic information.  Handling the purified DNA will generally

require less stringent precautions than will propagating the DNA. However,

the DNA itself should be handled at least as carefully as one would handle

the most dangerous of the DNAs used to make it.

  The above rule by itself effectively precludes certain experiments--

namely, those in which one of the components is in Class 5 of the

"Classification of Etiologic Agents on the Basis of Hazard" (5), as these

are excluded from the United States by law and USDA administrative policy.

There are additional experiments which may engender such serious biohazards
that they are not to be performed at this time. The&e are considered prior

to presentation of the containment guidelines for permissible experiments.

  A. Experiments that are not to be performed - We recognize that it
can be argued that certain of the recombinants placed in this category

could be adequately contained at this time.  Nonetheless, our estimates

of the possible dangers that may ensue if that containment fails are of


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such a magnitude that we consider it the wisest policy to at least defer

experiments on these recombinant DNAs until there is more information to

accurately assess'that danger and to allow the construction of more effective

biological barriers.  In this respect, these guidelines are more stringent
than those initially recommended (1).

  The following experiments are not to be initiated at the present time:
(1') Cloning of recombinant DNAs derived from the pathogenic organisms in
Classes 3, 4, and 5 of "Classification of Etiologic Agents on the Basis of

Hazard" (5), or oncogenic viruses classified by NC1 as moderate risk (6), or

cells known to be infected with such agents, regardless of the host-vector
system used. (ii) Deliberate formation of recombinant DNAs containing genes

for the biosynthesis of potent toxins (e.g., botulinum or diphtheria toxins;

venoms from insects, snakes, etc.).  (iii) Deliberate creation from plant

pathogens of recombinant DNAs that are likely to increase virulence and
host range. (iv) Deliberate release into the environment of any organism

containing a recombinant DNA molecule.  (v) Transfer of a drug resistance
trait to microorganisms that are not known to acquire it naturally if such
acquisition could compromise the use of a drug to control disease agents in
human or veterinary med'icine or agriculture.

  In addition, at this time large-scale experiments (e.g., more than 10
liters of culture) with recombinant DNAs known to make harmful products are not

to be carried out,  We differentiate between small- and large-scale experiments


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with such DNAs because the probability of escape from containment barriers

normally increases with increasing scale.  However, specific experiments in

this category that are of direct societal benefit may be excepted from this

rule if special biological containment precautions and

for large-scale operations are used, and provided that

are expressly approved by the Recombinant DNA Molecule

Committee of NIH.

equipment designed

these experiments

Program Advisory

  B. Containment guidelines for permissible experiments - It is an-

ticipated that most recombinant DNA experiments initiated before these
guidelines are next reviewed (i.e., within the year) will employ L. coli

K-12 host-vector systems.  These are also the systems for which we have

the most experience and knowledge regarding the effectiveness of the

containment provided by existing hosts and vectors necessary for the con-

struction of more effective biological barriers.
  For these reasons, E-, coli K-12 appears to be the system of choice

at this time, although we have carefully considered arguments that many

of the potential dangers are compounded by using an organism as intimately

connected with a man as is E. coli.  Thus, while proceeding cautiously
with E. coli, serious efforts should be made toward developing alternate
host-vector' systems; this subject is discussed in considerable detail in

Appendix A.

  We therefore consider DNA recombinants in F. coli K-12 before pro-

ceeding to other host-vector systems.