APRIL 27-28-MINUTES OF MEETING 
22 
Dr. Chakrabarty is interested in conducting two types of experiments. 
In the first, he wants to clone P. putida genes in P. putida using 
EL putida plaanids as vectors. There would be two kinds of vectors, 
PR4 1, a nonconj ugative plasmid, and RPl, a conjugative plasmid. 
In the second experiment. Dr. Chakrabarty wants to substitute 
EL coli or pMB9 DNA for P. putida DNA as a donor. Dr. Chakrabarty 
suggests Pi containment when the nonconj ugative plasmid is used 
and P2 when the conjugative plasmid is employed. 
Ihe RAC agreed that, according to the 1976 Guidelines, shotgun 
cloning of EL putida DNA into EL coli K-12 can be conducted under 
Pl-EKl conditions or even with conjugative plaanids under Pi 
conditions. 
The RAC felt that using EL putida as a host for EL coli DNA is 
not permitted under the 1976 Guidelines because of the require- 
ments that the host must be comparable to EKl in biological 
containment, i.e., free from conjugative plaanids. However, the 
RAC voted 9 to 1 to recommend that an exemption be granted to 
allow all of the requested experiments because EL putida and 
E. coli exchange ENA, and such pairs of organisms have been 
recommended for total exemption under the proposed revised 
Guidelines. 
The RAC was then specifically asked to judge which of the proposed 
experiments could be approved under the 1976 Guidelines. The Com- 
mittee noted that to be comparable to EKl the EL putida strains 
would have to be enfeebled and free of conjugative plasmids. 
The RAC then unanimously passed a motion to the effect that the 
investigators should be informed that, under the 1976 Guidelines, 
approval can be given under Pi conditions for those experiments 
in which no conjugative plasmids are present in the EL putida 
host when recombinant DNA is present, and after data have been 
submitted on (the poorer) survival in soil of the EL putida host 
strain compared to wild type. If these two conditions cannot be 
met, the RAC recommends that an exemption to the 1976 Guidelines 
be granted on the grounds that EL putida and Eh coli exchange 
DNA, and that these experiments have been recommended for total 
exemption under the revised Guidelines. 
C. In vitro Manipulation of Reccmbinant DNA Molecules 
The RAC reviewed a statement on the jri vitro handling of 
reccmbinant DNA prepared by the Harvard University Committee on 
the Regulation of Hazardous Biological Agents. The statement 
establishes different requirements for the vitro handling 
of recombinant ENA depending on whether the jri vivo requirements 
are Pl-EKl, or whether the _in vivo conditions require P2 or P3 
containment when in an EKl or EK2 host. 
[ 538 ] 
