Federal Register / Vol. 48. No. 106 / Wednesday, June 1, 1983 / Notices 
24519 
The exact wording of parts of 
Appendix L below was developed by 
NIH staff subsequent to the RAC 
meeting based on the recommendations 
made at the RAC meeting. The United 
States Department of Agriculture 
(USDA) Recombinant DNA Committee 
has reviewed the RAC recommendation, 
including the proposed wording for 
Appendix L given below, and has 
endorsed their adoption. 
I accept the recommendations of the 
RAC and the USDA Recombinant DNA 
Committee, and the following changes 
are incorporated into the Guidelines: 
Section III-A-2 would be amended to 
read as follows: 
Deliberate release into the environment of 
any organism containing recombinant DNA, 
except certain plants as described in 
Appendix L. 
A new appendix, Appendix L, would 
be added as follows: 
Appendix I.. Release Into the Environment 
of Certain Plants. 
Appendix L-I. General Information. 
Appendix L specifies conditions under 
which certain plants, as specified below, may 
be approved for release into the environment. 
Experiments in this category cannot be 
initiated without submission of relevant 
information on the proposed experiment to 
NIM, review by the RAC Plant Working 
Croup, and specific approval by NIH. Such 
experiments also require the approval of the 
IBC before initiation. Information on specific 
experiments which have been approved will 
be available in ORDA and will be listed in 
Appendix L-I1I when the Guidelines are 
republished. 
Experiments which do not meet the 
specifications of Appendix L-1I fall under 
Section III- A and require full RAC review 
and NIH and IBC approval before initiation. 
Appendix L-II. Criteria Allowing Review 
by the RAC Plant Working Croup Without 
The Requirement for Full RAC Review . 
Approval may be granted by ORDA In 
consultation with the RAC Plant Working 
Group without the requirement for full RAC 
review (IBC review is also necessary) for 
growing plants containing recombinant DNA 
in the field under the following conditions: 
Appendix L-II-A. The plant species is a 
cultivated crop of a genus that has no species 
known to be a noxious weed. 
Appendix L-II-B. The introduced DNA 
consists of well-characterized genes 
containing no sequences harmful to humans, 
animals, or plants. 
Appendix L-II -C. The vector consists of 
DNA (i) from exempt host-vector systems 
(Appendix C); (ii) from plants of the same or 
closely related species; (iii) from 
nonpalhogenic prokaryotes or nonpathogenic 
lower eukaryotic plants; (iv) from plant 
pathogens only if sequences causing disease 
have been deleted, or (v) chimeric vectors 
constructed from sequences defined in (i) to 
(iv) above. The DNA may be introduced by 
any suitable method. 
Appendix L-ll-D. Plants are grown in 
controlled access fields under specified 
conditions appropriate for the plant under 
study and the geographical location. Such 
conditions should include provisions for 
using good cultural and pest control 
practices, for physical isolation from plants of 
the same species outside of the experimental 
plot in accordance with pollination 
characteristics of the species, and for 
preventing plants containing recombinant 
DNA from becoming established in the 
environment. Review by the IBC should 
include an appraisal by scientists 
knowledgeable of the crop, its production 
practices, and the local geographical 
conditions. Procedures for assessing 
alternations in and the spread of organisms 
containing recombinant DNA must be 
developed. The results of the outlined test 
most be submitted to and reviewed by the 
IBC. Copies must also be submitted to the 
RAC Plant Working Croup. 
Appendix L-lll. Specific Approvals. 
B. Release of Strains of Pseudomonas 
Syringae and Erwinia Herbicola. 
Drs. Nickolas Panopoulos and Steven 
l.indow of the University of California, 
Berkeley, submitted a revised request to 
construct and release Pseudomonas 
syringae pv. syringae and Erwinia 
herbicola carrying in vitro generated 
deletions of all or part of the genes 
involved in ice nucleation for purposes 
of biological control of frost damage in 
plants. The aim of the experiments is to 
investigate possibilities for biological 
control of frost damage in plants. 
Certain bacteria, such as 
Pseudomonas syringae and Erwinia 
herbicola cause nucleation of ice 
crystals. These bacteria are common 
plant epiphytes. A causal relationship 
has been established between frost 
damage on forst-senstive crop plants in 
the temperature range of 0° to — 5°C, 
and the populations of ice nucleation 
active bacteria present on the plants. 
There are chemically induced mutants 
of P. syringae and E. herbicola that do 
not cause ice nucleation, and their 
effectiveness under field conditions has 
been demonstrated. The investigators 
now propose to construct deletion 
mutants for ice nucleation activity in 
prototype strains of P. syringae and E. 
herbicola and to evaluate their efficacy 
as bilogical competitors of naturally 
occuring populations of these bacteria. 
The investigators state that the 
advantages of such mutants compared 
to chemically induced equivalents are 
genetic stability and the absence of 
silent mutations which may adversely 
effect competitive fitness. Prior to the 
field applications, the investigators plan 
to test the mutant strains in a contained 
environment, such as growth chambers 
and greenhouse, to verify that they do 
not induce frost or other injury to plants, 
and that they are capable of colonizing 
leaves. 
The RAC reviewed a similar proposal 
from these investigators at its October 
25, 1982, meeting. The RAC had passed a 
motion recommending approval of the 
requested field tests. However, approval 
by NIH was deferred because of 
concerns raised at that meeting (48 FR 
1158). 
The Revised proposal was 
summarized in the March 4, 1983, 
Federal Register (48FR 9441). After the 
comment period, USDA received one 
letter urging the RAC to consider the 
request favorably. 
The RAC reviewed the revised 
proposal at the April 11. 1983, meeting. It 
was pointed out that the revised 
proposal responds to the concerns 
previously raised. The investigators 
propose to use chromosomal antibiotic 
resistance markers to monitor the 
strains. The revised proposal discusses 
several antibiotics not used in clinical 
medicine that could be tried, in addition 
to refamycin, for marking of the strains 
with chromosomal antibiotic resistance. 
The revised submission to conduct field 
testing at a single location, the 
University of California Field Station at 
Tulelake in Northern California, rather 
than six locations requested previously. 
The investigators noted that this site is 
geographically isolated from all major 
fruit tree and citrus growing regions of 
the state. It was pointed out at the RAC 
meeting that fields have already been 
sprayed with chemically induced 
(nonrecombinant DNA) mutant 
organisms. Also, it was noted that the 
populations of bacteria proposed to be 
released are approximately eight to ten 
orders of magnitude lower than you 
would find normally at any given time of 
year. With regard to an emergency plan 
described in the proposal, the RAC did 
not recommend spraying with antibotics 
to eliminate organisms; they 
recommended instead either burning or 
preferably burying the material. The 
RAC concluded that the investigators 
had satisfactorily answered the issues 
raised at the previous meeting and voted 
to recommend approval of the proposal 
excluding the use of antibiotics in 
emergency procedures. The vote was 
nineteen in favor, none opposed, and no 
abstentions. 
The USDA Recombinant DNA 
Committee reviewed the proposal and 
recommended that it be approved. 
I accept the recommendations of the 
RAC and the USDA Recombinant DNA 
Committee, and the following wording 
has been added to Appendix D: 
Appendix D-X. Permission is granted to 
Drs. Steven Lindow and Nickolas Panopoulos 
of the University of California, Berkeley, to 
release under specified conditions 
ri^oi 
