9 
"a) remove from the original containment conditions any clones of 
fragments of the phage genome (generated during the process of 
obtaining cloned toxin structural genes) that do not correspond 
to toxin structural genes and to work with them under PI EK1 
conditions; and 
"b) directly clone any fragments of the phage gencme that do not 
correspond to toxin structural genes into coli K-12 under PI 
EK1 conditions." 
It was the consensus of the group that these experiments would not fall under 
Appendix F of the Guidelines, and no action need be taken. 
Dr. Gottesman requested the group to consider the fifth item of Dr. O'Brien's 
proposal. That item reads as follows: 
"5. If in future experiments we can isolate nontoxinogenic alleles of 
the structural gene(s) for Shiga-like toxin by transpo son-media ted 
mutagenesis (insertional inactivation) or by chemical mutagensis, we 
request permission to clone these nontoxinogenic alleles of the toxin 
structural gene(s) into Eh_ coli K-12 under PI + EK1 conditions." 
It was the consensus of the group that because no working group could predict 
all potential scenarios each specific nontoxinogenic allele should be considered 
individually on a case-by-case basis. A system is in place within the NIH to 
perform this type of evaluation. 
Dr. Gottesman asked if the language or the philosophical basis of Appendix F 
should be modified in light of the working group discussion concerning the 
cloning of the Shiga toxin gene. The working group believed that no modifications 
were required in Appendix F. 
Dr. Gottesman adjourned the meeting of the Working Group on Tbxins at 11:30 a.m.. 
May 11, 1984. 
[ 418 ] 
