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Federal Register / Vol. 44, No. 232 / Friday, November 30, 1979 / Notices 
systems for HV3 certification will be 
performed by contractors selected by 
NIH. These contractors will repeat tests 
performed by individuals proposing the 
HV2 system and, in addition, will 
conduct more extensive tests on 
conditions likely to be encountered in 
nature. The genotypic and phenotypic 
traits of HV2 systems will be evaluated. 
Tests on survival and transmissibility in 
and on animals, including primates, will 
be performed, as well as tests on 
survival in certain specified natural 
environments. 
II-D-3. Distribution of -Certified Host- 
Vectors. Certified HV2 and HV3 host- 
vector systems (plus appropriate control 
strains) must be obtained from the NIH 
or its designees, one of whom will be the 
investigator who developed the system. 
NIH shall announce the availability of 
the system by publication of notices in 
appropriate journals. 
Plasmid vectors will be provided in a 
suitable host strain, and phage vectors 
will be distributed as small-volume 
lysates. If NIH propagates any of the 
host strains or phage, a sample will be 
sent to the investigator who developed 
the system or to an appropriate 
contractor, prior to distribution, for 
verification that the material is free from 
contamination and unchanged in 
phenotypic properties. 
In distributing the certified HV2 and 
HV3 host-vector systems, NIH or its 
designee will (i) send out a complete 
description of the system; (ii) enumerate 
and describe the tests to be performed 
by the user in order to verify important 
phenotypic traits; (iii) remind the user 
that any modification of the system 
necessitates independent approval of 
the system by the NIH; and (iv) remind 
the user of responsibility for notifying 
ORDA of any discrepancies with the 
reported properties or any problems in 
the safe use of the system. 
NIH may also distribute certified HVl 
host-vector systems. 
III. Containment Guidelines for Covered 
Experiments 
Part III discusses experiments covered 
by the Guidelines. The reader must first 
consult Part I, where listings are given of 
prohibited and exempt experiments. 
Containment guidelines for 
permissible experiments are given in 
Part III. Changes in these levels for 
specific experiments (or the assignment 
of levels to experiments not explicitly 
considered here) may not be instituted 
without the express approval of the 
Director, NIH. (See Sections IV-E-l-b- 
(l)-(a), IV-E-l-b-(l)-(b), IV-E-l-b-(2}- 
(b), IV-E-l-b-(2)-(c), and IV-E-l-b-(3)- 
(b)0 
In the following classification of 
containment criteria for different kinds 
of recombinant DNAs, the stated levels 
of physical and biological containment 
are minimal for the experiments 
designated. The use of higher levels of 
biological containment 
(HV3>HV2>HV1) is encouraged if they 
are available and equally appropriate 
for the purposes of the experiment. 
Ill— 0. Classification of Experiments 
Using the E. coli K-12 Host- Vector 
Systems. Most recombinant DNA 
experiments currently being done 
employ E. coli K-12 host-vector systems. 
These are the systems for which we 
have the most experience and 
knowledge. 
Some experiments using E. coli K-12 
host-vector systems are prohibited (see 
Section I-D). 
Some experiments using E. coli K-12 
host-vector systems are exempt from the 
Guidelines (see Section I-E). 
Other experiments using E. coli K-12 
shall use Pi physical containment and, 
except as specified in the last paragraph 
of this section, an EK1 host-vector 
system (i.e. (a) the host shall not contain 
conjugation-proficient plasmids or 
generalized transducing phages, and (b) 
lambda or lambdoid bacteriophages or 
non-conjugative plasmids shall be used 
as vectors). For these experiments no 
Memorandum of Understanding and 
Agreement (MUA) as described in 
Section IV-D-l-c need be submitted, 
nor is any registration with NIH 
necessary. However, for these 
experiments, prior to their initiation, 
investigators must submit to their 
Institutional Biosafety Committee (IBC) 
a registration document that contains a 
description of (a) the source(s) of DNA, 
(b) the nature of the inserted DNA 
sequences, and (c) the hosts and vectors 
to be used. This registration document 
must be dated and signed by the 
investigator and filed only with the local 
IBC. The IBC shall review all such 
proposals but such review is not 
required prior to initiation of 
experiments. An exception, however, 
which does require prior review and 
approval by the IBC is any experiment 
in which there is a deliberate attempt to 
have the E. coli K-12 efficiently express 
any gene coding for a eukaryotic 
protein. 
Experiments involving the insertion 
into E. coli K-12 of DNA from 
prokaryotes that exchange genetic 
information with E. coli by known 
physiological processes will be 
exempted from these Guidelines if they 
appear on the "list of exchangers" set 
forth in Appendix A (see Section I— E — 4). 
For those not on the Appendix A list 
but which exchange genetic information 
[ 204 ] 
[35] with E. coli, experiments may be 
performed with any E. coli K-12 vector 
(e.g. conjugative plasmid). When a non- 
conjugative vector is used, the E. coli K- 
12 host may contain conjugation- 
proficient plasmids, either autonomous 
or integrated, or generalized transducing 
phages. 
Ill— A. Classification of Experiments 
Using Certain HVl and HV2 Host- 
Vector Systems. Certain HVl and HV2 
host-vector systems are assigned 
containment levels as specified in the 
subsections of this Section III— A. Those 
so classified as of publication of these 
revised Guidelines are listed in 
Appendix D. An updated list may be 
obtained from the Office of 
Recombinant DNA Activities, National 
Institutes of Health, Bethesda, Maryland 
20205. 
It has been necessary, throughout this 
section, to use Words and terms marked 
with footnote reference numbers. The 
footnootes (Part V) define more fully 
what the terms denote. 
III-A-4. Shotgun Experiments. These 
experiments involve the production of 
recombinant DNAs between the vector 
and portions of the specified cellular 
source, preferably a partially purified 
fraction. Care should be taken either to 
preclude or eliminate contaminating 
micro-organisms before isolating the 
DNA. 
III-A-l-a. Eukaryotic DNA 
Recombinants. 
III-A-l-a-(l). Primates. P2 physical 
containment + an HV2 host-vector or 
P3 + HVl. 
III-A-l-a-(2). Other Mammals. P2 
physical containment -|- an HV2 host- 
vector or P3 + HVl. 
Ill— A— 1— a— (3). Birds. P2 physical 
containment + an HV2 host-vector, or 
P3 + HVl. 
III-A-l-a-(4). Cold-Blooded 
Vertebrates. P2 physical containment + 
an HVl host-vector or Pi + HV2. If the 
eukaryote is known to produce a potent 
polypeptide toxin, [34] the containment 
shall be increased to P3 + HV2. 
III-A-l-a-(5). Other Cold-Blooded 
Animals and Lower Eukaryotes. This 
large class of eukaryotes is divided into 
two groups: 
III-A-l-a-(5)-(a). Species that are 
known to produce a potent polypeptide 
toxin [34] that acts in vertebrates, or are 
known pathogens listed in Class 2, [1] or 
are known to carry such pathogens must 
use P3 physical containment + an HV2 
host-vector. When the potent toxin is 
not a polypeptide and is likely not to be 
the product of closely linked eukaryote 
genes, containment may be reduced to 
P3 + HVl or P2 + HV2. Species that 
produce potent toxins that affect 
invertebrates or plants but not 
