7 
Dr. Gill said he would like Dr. Levine's motion to require the investigators to 
select organisms with lower levels of toxin expression than 10 7 50% cytotoxic 
doses/mg protein in cell lysates and 10 6 50% cytotoxic doses/ml in culture 
supernatants. 
Dr. Formal said an organism producing 10 7 50% cytotoxic doses of toxin would 
be no different than those currently in nature. Dr. Gottesman said the 50% 
cytotoxic dose measurement had been made under laboratory growth conditions. 
She wondered whether the modified organism might express different levels of 
toxin under laboratory growth conditions than in nature. 
Dr. Levine pointed out that use of poorly mobilizable plasmid vectors such as 
pBR322 add an additional measure of safety; language specifying use of this 
type of plasmid vector might meet Dr. Gill's concerns. Dr. Holmes pointed out 
that the structural gene for Shiga toxin may be on a phage and may be widely 
disseminated in nature. 
Dr. Gill offered a substitute motion for Dr. Levine's motion. Dr. Gill's 
substitute motion read as follows: 
"The working group recommends to RAC that the second item in Dr. O'Brien's 
request be permitted under the following conditions: 
"(1) containment would be set at P2; 
"(2) poorly nobilizable plasmids such as pBR322, pBR325, and pBR328 
would be used as vectors; and 
"(3) the organisms removed from P3 containment would produce ten fold 
less toxin than highly toxinogenic clinical isolates of EL_ coli 
(grown in glucose syncase media)." 
Dr. Levine said his original motion as amended reads much like Dr. Gill's 
substitute motion except that he would "prefer" rather than "require" that the 
clones to be removed from P3 containment be constructed using poorly mobilizable 
plasmids such as pBR322 or pBR328. 
In order to permit a vote. Dr. Formal seconded Dr. Gill's substitute motion. 
By a vote of two in favor, three opposed, and no abstentions, Dr. Gill's substi- 
tute motion failed. Dr. Levine's motion thus became the motion to be considered. 
Dr. Levine's motion reads as follows: 
"The working group recommends to RAC that the request to remove Shiga toxin 
clones from P3 containment be granted under the following conditions: 
"(1) containment would be P2; 
"(2) the level of expression of toxin by the organism would be ten 
fold less than toxin expression by clinical isolates (i.e., 10^ 50% 
[416] 
