37 
that mouse cells, derived preferably from embryos, be 
used as the source of eukaryotic DNA. Polyoma virus 
is a mouse virus and recombinant DNA molecules contain- 
ing both viral and cellular sequences are already known 
to be present in virus stocks grown at a high multiplic- 
ity. Thus, recombinants formed in vitro between polyoma 
virus DNA and mouse DNA are presumably not novel from 
an evolutionary point of view. 
b_. Such experiments can be done, under P4 conditions, if the 
recombinant DNA contains segments of the genomes of Class 
2 animal viruses (5). Once it has been shown by suit- 
able biochemical and biological tests that the cloned 
recombinant contains only harmless regions of the viral 
genome 'see Section IIIB-2-c-i) and that the host range 
of the polyoma virus vector has not been altered, ex- 
periments can be continued under P3 conditions. 
2. SV40 Virus 
a_. Defective SV40 genomes, with appropriate helper, can 
be used in P4 conditions as a vector for recombinant DNA 
mo'ecules containing sequences of any non-pathogenic 
organism or Class I virus (5), (i.e., a shotgun type 
experiment); established lines of cultured cells should 
be used. 
b_. Such experiments can be carried out in P3 conditions if 
the non-SV40 DNA segment is (a) a purified' segment of 
prokaryotic DNA lacking toxigenic genes, or (b) a segment 
g 
1 he Jl\ A preparation is defined as purified if the desired DNA represents 
at _east 99% (w/w) of the total DNA in the preparation, provided that it 
was achieved or verified by more than one procedure. 
[108] 
