Recombinant DNA Advisory Committee - 09/14-15/92 
or endogenous protease activity that activates the virus, or (3) a genetic revertant. In the 
case of the SQL mutation, the estimated frequency of reversion is approximately 10' 12 . It 
should be noted that a second site revertant would probably occur at a higher frequency. 
Dr. Temple presented plaque assay data demonstrating that when equal numbers of 
infectious units of the Helper 1 package virion and cells (10 6 ) were combined, 
approximately 700 to 10,000 plaque forming units were observed. This corresponds to 
the number of units that would result from one or two recombinant viruses. Therefore, 
1 in 10 6 recombinant particles are detectable using this assay. Additional plaque assays 
demonstrated no infectious particles using the SQL helper virus. In fact, recombination 
plus suppression of the SQL effect was required for the generation of replication 
competent virus. He showed in vivo experiments in which 24 newborn and 40-day-old 
mice received intracerebral or intranasal injections of greater than 10 8 infectious units of 
the packaged SQL helper virus. No evidence of replication competent virus was 
demonstrated. 
Dr. Temple stated that earlier data indicated that the recombination frequency of this 
virus is approximately KT 6 , and the frequency of "leakiness" is approximately 1CT 6 . 
However, both events would have to occur in order for replication competent virus to 
emerge. Therefore, even if the kit is used under circumstances other than those 
specifically outlined, there is a wide margin of safety. Because this is an efficient system, 
it is in high demand by many researchers. It is very important that its use be approved 
at the BL2 level of physical containment. Researchers using this system at the BL2 level 
will be required to sign an acknowledgement that they have been adequately informed of 
the procedures necessary to minimize replication competent virus emerging and that they 
are knowledgeable of BL2 requirements. An independent confirmation will also be 
required regarding the principal investigator's (Pi's) expertise and qualifications by the 
Institutional Biosafety Committee. In addition, the PI will be requested to sign an 
agreement that he/she will not distribute the kit to other investigators without prior 
approval. Although there is no guarantee that the investigator will adhere to these 
principles, this process provides a degree of safety. 
Discussion 
Dr. Schaechter explained that because this kit will not fall under FDA regulations, the 
RAC is in the rare situation of deciding the disposition of this material on an advisory 
basis, not a statutory basis. Dr. Murray asked the investigators to respond to Dr. 
Schaechter's concern regarding other regulatory bodies that would provide approval of 
this kit. Dr. Temple stated that there is no other regulatory body whose approval is 
required. 
Dr. Doi noted that the investigators stated that this kit was already in use by some 
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