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Federal Register / Vol. 46. No. 233 / Friday, December 4, 1981 / Notices 
Appendix C of Annex E. — Exemptions 
under 1-E-^ 
Section I-E-5 states that exempt from 
these Guidelines are "Other classes of 
recombinant DNA molecules, if the 
Director, NIH. with advice of the 
Recombinant DNA Advisory Committee, 
after appropnate notice and opportunity 
for public comment, finds that they do 
not present a significant risk to health or 
the environment. (See Section IV-E-1- 
b-(lHd).) Certain classes are exempt as 
of publication of these Revised 
Guidelines.” 
The following classes of experiments 
are exempt under Section 1-E^ of the 
Guidelines; 
1. Recombinant DNAs in Tissue 
Culture. 
Recombinant DNA molecules derived 
entirely from non-viral components (that 
is. no component is derived &om a 
eukaryotic virus), that are propagated 
and maintained in cells in tissue culture 
are exempt from these Guidelines with 
the exceptions listed below. 
Exceptions. 
Experiments described in Sections 1- 
D-1 to I-D-5 as being prohibited. 
Experiments involving DNA from 
Class 3 organisms (1] or cells known to 
be infected with these agents, or any 
recombinant DNA molecules which 
Increase the virulence and host-range of 
a plant pathogen beyond that which 
occurs by natural genetic exchange. (See 
Section IIl-O-l.) 
Experiments involving the deliberate 
introduction of genes coding for the 
biosynthesis of toxins potent for 
vertebrates. (See Appendix G.) 
2. Experiments Involving E coli K-12 
host-vector systems. 
Experiments which use E. coli K-12 
host-vector systems, with the exception 
of those experiments listed below, are 
exempt from these Guidelines provided 
that (a) the E. coli host shall not contain 
conjugation proficient plasmids or 
generalized transducing phages, and (b) 
lambda or lambdoid or Ff 
bacteriophages or nonconjugative 
plasmids (49| shall be used as vectors. 
However, experiments involving the 
insertion into E. coli K-12 of DNA from 
prokaryotes that exchange genetic 
information [35] with E. coli may be 
perforfned with any E. coli K-12 vector 
(e g., conjugative plasmid). When a 
nonconjugative vector is used, the E. 
coll K-12 host may contain conjugation- 
proHcient plasmids either autonomous 
or integrated, or generalized transducing 
phages. 
For these exempt experiments. Pi 
physical containment conditions are 
recommended. 
Exceptions. 
Experiments described in Sections I- 
D-1 to I-D-5 as being prohibited. 
Experiments involving DNA from 
Class 3 organisms [1] or from cells 
known to be infected with these agents 
may be conducted at P3 containment. 
Lower containment levels may be 
specified by NIH. (See Section IV-E-1- 
b^2)-(e).) Experiments in this category 
require prior IBC review and approval. 
Experiments which increase the 
virulence and host range of a plant 
pathogen beyond that which occurs by 
natural genetic exchange. (See Section 
UI-O-1.) 
Large-scale experiments (e g., more 
than 10 liters of culture) require prior 
IfiC review and approval. 
Experiments involving the deliberate 
cloning of genes coding for the 
biosynthesis of toxins potent for 
vertebrates. (See Appendix G.) 
3. Experiments Involving 
Saccharomyces cerevisiae host-vector 
systems. 
Experiments which use 
Saccharomyces cerevisiae host-vector 
systems, with the exception of 
experiments listed below, are exempt 
from these Guidelines provided that 
laboratory strains are used. 
For these exempt experiments. Pi 
physical containment conditions are 
recommended. 
Exceptions. 
Experiments described in Sections I- 
D-1 to I-D-5 as being prohibited. 
Experiments involving GDC Class 3 
organisms [1] or cells known to be 
Infected %vith these agents, or any 
recombinant DNA molecules which 
increase the virulence and host-range of 
a plant pathogen beyond that which 
occurs by natural genetic exchange. (See 
Section IIl-O-l.) 
Large-scale experiments (e.g., more 
than 10 Uters of culture) require prior 
IBC review and approval. 
Experiments involving the deliberate 
cloning of genes coding for the 
biosynthesis of toxins potent for 
vertebrates. (See Appendix G.) 
4. Experiments Involving Bacillus 
subtilis host-vector systems. 
Any asporogenic Bacillus subtilis 
strain which does not revert to a 
sporeformer with a frequency greater 
than 10* ’can be used for cloning DNA 
from any nonprohibited source, with the 
exception of those experiments listed 
below. Indigenous Bacillus plasmids 
and phages, whose host-range does not 
include Bacillus cereus or Bacillus 
anthracis, may be used as vectors. 
For these exempt experiments Pi 
physical containment conditions are 
recommended. 
Exceptions. 
Experiments described in Section I-D- 
1 to I-D-5 as being prohibited. 
Experiments involving GDC Class 3 
organisms [1] or cells known to be 
infected with these agents, or any 
recombinant DNA molecules which 
increase the virulence and host-range of 
a plant pathogen beyond that which 
occurs by natural genetic exchange. (See 
Section IIl-O-l.) 
Large-scale experiments (e.g., more 
than 10 liters of culture) require prior 
IBC review and approval. 
Experiments involving the deliberate 
cloning of genes coding for the 
biosynthesis of toxins potent for 
vertebrates. (See Appendix G.) 
Appendix D of Annex E. — HVl and HV2 
Host-Vector Systems Assigned 
Containment Levels as Specified in the 
Subsections of Section III-A 
As noted above at the beginning of 
Section Ill-A, certain HVl and HV2 
host-vector systems are assigned 
containment levels as specified in the 
subsections of Section III-A. Those so 
classified as of publication of these 
Revised Guidelines are listed below. 
*HVl — The following specified strains 
of Neurospora crassa which have been 
modified to prevent aerial dispersion; 
(1) ini (inositolless) strains 37102, 
37401, 46316. 64001 and 89601. 
(2) csp-1 strain UCLA37 and csp-2 
strains FS 590, UCLAlOl (these are 
conidial separation mutants). 
(3) eas strain UCLAlOl (an "easily 
weltable” mutant). 
HVl — The foWov/ing Streptomyces 
species; Streptomyces coelicolor, S. 
lividans, S. porvulus. and S. griseus. The 
following are accepted as vector 
components of certified Streptomyces 
HVl systems; Streptomyces plasmids 
SCP2. SLPl.2, pl)101, actinophage phi 
C31, and their derivatives. 
Appendix E of Annex E. — Actions 
Taken Under the Guidelines 
As noted in (he subsections of 
Sections IV-E-l-b-(l) and IV-E-l-b-(2), 
the Director, NIH, may take certain 
actions with regard to the Guidelines 
after consideration by the RAC. 
Some of the actions taken to date 
include the following; 
1. The following experiment has been 
approved: The cloning in B. subtilis, under P2 
conditions, of DNA derived from 
Saccharomyces cerevisiae using EK2 plasmid 
*Thei« follow Ihe assigned conlainmeni levels as 
specified in Ihe subsectiona of Seclion III-A with 
one exception. This exception is that experiments 
involving complete genomes of eukaryotic viruses 
will require P3 + HV1 or P2 + HV2 rather than the 
levels given in the subscclinns of Section III-A. 
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I 
