Fig 5. SI nuclease analysis of NIH 3T3 cells Infected with N2 and N4 virus. 
SI nuclease analysis of total cytoplasmic RNA was performed according to Berk 
and Sharp (1) as modified by Weaver and Weissman (18). The presence and 
position of a 3 ' splice site (or the 5' end of a novel RNA species) was 
determined by using a probe spanning the junction of Neo and upstream viral 
sequences. The DNA probe was labeled at the Ncol site 599 bp within the Neo 
sequences extending to a Pvul site within the viral sequences 279 bp downstream 
from the LTR. For the N4 vector, the size of the probe was approximately 650 
bp, and for the N2 vector, the size was exactly 1220 bp (the junction of N2 but 
not N4 was determined by DNA sequencing). In each case (lanes N4 and N2) 
hybridization of the cytoplasmic RNA obtained from N4 or N2 infected NIH 3T3 
cells led to the formation of one SI nuclease resistant band which comigrated’ 
wit the corresponding probe, not present when RNA from uninfected NIH 3T3 cells 
was used (not shown). 
Recombinant DNA Research, Volume 12 
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