Federal Register / Vol. 45, No, 164 / Thursday, August 21, 1980 / Notices 
55925 
carrying Arabidopsis DNA and cloned 
in E. coli K-12. 
(d) The hybrid plasmid into which 
Arabidopsis DNA and the thiamine gene 
have been ligated will be transformed 
into Agrobacterium tumefaciens. 
(e) Agrobacterium tumefaciens will be 
used to introduce the E. coli / Ti plasmid 
vector carrying the E. coli thiamine gene 
and Arabidopsis DNA into Arabidopsis 
plants. 
According to Section III— O of the 
Guidelines, steps, (a), (b) and (c) of the 
protocol may be performed under Pi 
containment; according to Appendix E, 
steps (d) and (e) may be conducted 
under P3 containment. Dr. Merlo 
requests permission to perform steps (d) 
and (e) at Pi or P2 containment. He 
suggests that the DNA combination to 
be generated could occur in nature, 
albeit at very low frequency. 
4. Request for certification of a 
Bacillus subtilis strain as the host- 
vector component of an HV2 host-vector 
system. On March 28, 1980, Dr. William 
F. Burke, Jr.,' of Arizona State University, 
requested certification of Bacillus 
subtilis strain ASB298 as the host 
component of an HV2 host-vector 
system. Dr. Burke’s proposal was 
evaluated by a working group and 
subsequently presented to the RAC at 
the June 5-6, 1980 meeting. The RAC 
discussed the characteristics of a 
Bacillus subtilis HV2 system and 
recommended that consideration of the 
proposal be deferred until additional 
information concerning transfer of 
recombinant DNA through 
transformation was received. The RAC 
will continue the evaluation of 
additional information received on 
strain ASB298 at the September meeting. 
5. Request for HVl certification of a 
Bacillus stearothermophilus derived ' 
plasmid vector in Bacillus subtilis. Dr. 
David B. Wilson of Cornell University, 
requests HVl certification of a host- 
vector system based on a Bacillus 
subtilis host and a plasmid isolated from 
Bacillus stearothermophilus. 
6. Proposed revision of subsections of 
section III-C-1-e. A notice appeared in 
the Federal Register of January 31, 1980 
concerning proposed revision of Section 
III-C-1-e, and its subsections. It was 
recommended that Sections II-C-1-e, 
Ill_c-i_e_{i), III— C— 1 — e— ( 1 )— (a), and III- 
C— 1— e— (1)— (b), of the Guidelines be 
changed and that a new Section III— C— 1— 
e-(l)-(c) be added. Section III-C-l-e-{2) 
would remain unchanged. The RAC, at 
its March 6-7, 1980 meeting, 
recommended adoption of Sections HI— 
C-l-e, UI-C-l-e-{l), and IU-C-e-(lHa) 
as published in the Federal Register of 
January 31, 1980, with certain 
modifications in Section III— C— 1— e— (1 )— 
(a). 
The Director, NIH, accepted this 
recommendation and promulgated the 
following sections in the Federal 
Register of April 14, 1980: 
"III-C-1-e. All Viral Vectors. 
HI — C— 1— e — (lj. Other experiments 
involving eukaryotic virus vectors can 
be done as follows: 
III-C-l-e-(lHa)- Recombinant DNA 
molecules containing no more than two- 
thirds of the genome of any eukaryotic 
virus [all viruses form a single Family 
(36) being considered identical (50)] may 
be propagated and maintained in cells in 
tissue culture using Pi containment. For 
such experiments, it must be shown that 
the cells lack helper virus for the 
specific Families of defective viruses 
being used. The DNA may contain 
fragment of the genomes of viruses from 
more than one Family but each fragment 
must be less than two-thirds of a 
genome.” 
At its March 6-7, 1980 meeting, the 
RAC voted to defer consideration until 
the June 5-6, 1980 meeting of the new 
Sections III— C— 1— e(l)— fb) and UI-C-1-e- 
(l)-(c) as proposed in the Federal 
Register of January 31, ,1980, and 
requested that a working group develop 
additional information. Accordingly, a 
working group met on May 13, 1980 
during the annual meeting of the 
American Society for Microbiology in 
Miama Beach, Florida. The report of the 
working group was considered briefly at 
the June 5-6 RAC meeting and will be 
considered again by the RAC at its 
September 25-26, 1980 meeting. 
The Working Group discussed the 
question of the appropriate containment 
conditions for experiments involving 
recombinant DNA molecules containing 
less than two-thirds of the genome of 
any eukaryotic virus which may be „ 
rescued with helper virus. On the basis 
of the consensus of the virologists, the 
following recommendation was 
proposed, as a revision of Section III-C- 
l-e-(l)-(b) of the Guidelines 
“UI-C-l-e-(l)-(b). Recombinants with 
less than two-thirds of the genome of 
any eukaryotic virus may be rescued 
with helper virus using P2 containment 
if wild type strains of the virus are CDC 
Class 1 or 2 agents, or using P3 
containment if wild type strains of the 
virus are CDC Class 3 agents (1).” 
7. Procedures for review of large-scale 
experiments. Section I-D-6 of the 
Guidelines prohibits "large-scale 
experiments (e.g., more than 10 liters of 
culture) with organisms containing 
recombinant DNAs, unless the 
recombinant DNAs are rigorously 
characterized and the absence of 
harmful sequences established." 
Section IV-E-l-b-(3)-(d) of the 
Guidelines states that the Director, NIH, 
is responsible for “authorizing, under 
procedures specified by the RAC, large- 
scale experiments (i.e., more than 10 
liters of culture) for recombinant DNAs 
that are rigorously characterized and 
free of harmful sequences.” 
Part VI of the Guidelines, "Voluntary 
Compliance,” encourages institutions 
not otherwise covered by the Guidelines 
to follow the standards and procedures 
set forth in the Guidelines. 
At its September 1979 meeting, the 
RAC adopted the following procedures 
to be followed by applicants proposing 
to exceed the 10-liter limit: 
"Application Procedures For Large- 
Scale Recombinant DNA Experiments 
1. For each research project proposing 
to exceed the 10-liter limit, the applicant 
shall file a request with the NIH Office 
of Recombinant DNA Activities 
(ORDA). The request should include the 
following information: 
a. The Memorandum of 
Understanding and Agreement (MUA) 
submitted to the local Institutional 
Biosafety Committee. The MUA should 
include, or have appended to it, a 
summary paragraph which describes the 
proposed project in language that is 
comprehensible to non-specialists. 
b. A statement of the rationale for 
wishing to exceed the 10-liter limit. 
c. A specification of the total volume 
of the fermenter to be used. 
d. Evidence that the recombinant 
DNAs to be employed in the research 
have been rigorously characterized and 
are free of harmful sequences. 
e. A description of the applicant’s 
laboratory practices, containment 
equipment, and facilities relevant to the 
containment of large volumes of culture. 
f. Evidence of the applicant’s or 
applicant institution’s previous 
experience in handling large volumes of 
culture. Applicants should exhibit 
knowledge of state-of-the-art procedures 
for working with large volumes of 
microorganisms. 
g. A, description of procedures to be 
employed for the inactivation and 
disposal of large volumes of culture. 
h. A description of procedures for 
containing and inactivating accidental 
spills, should they occur. 
2. Each request submitted to ORDA 
shall be referred to a working group of 
the NIH Recombinant DNA Advisory 
Committee for review. 
3. Following review and approval by 
the working group, each request shall be 
submitted to the entire Recombinant 
DNA Advisory Committee for review. 
4. Following review and approval by 
the RAC, each request shall be 
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