Characterization of poly(adenosine diphosphate ribosylated) protein isolated from mouse L1210 cells in vivo [proceedings].

dence from Schell et al (1979), who investigated the loss of phenotypic characters resulting from the insertion of the transposon Tn7 into Ti plasmids, which suggests that the nopaline synthetic genes, which will show, at best, low homology with plasmid-pTiB6S3 sequences, should be located close to the end of the T-DNA. Although it has been shown that Escherichia coli RNA polymerase in vitro will transcribe portions of the T-DNA (Drummond et al., 1977; Ledeboer, 1978), maxicells (Sancar et al., 1979) containing plasmid pSZ595 make plasmid-encoded proteins only from vector-pMB9 genes. We have investigated the maxicell-RNA transcripts of this plasmid to determine whether the apparent block to expression in vivo occurs a t transcription or at translation. Subfractionation of the endonuclease-HindIII-produced fragment containing the end of the T-DNA has proved difficult, as few restriction endonucleases cleave in this region. In addition to those enzymes shown in Fig. 1, endonucleases HgiAI and HaeIII all generate large (greater than 0.9 x lo6 daltons) fragments in this region. We have been screening enzymes whose recognition site comprises four bases in order to break up this region into lengths suitable for sequencing. One piece of gross sequence information obtained to date is that, at a distance corresponding to 3 x 106-4 x lo6 daltons to the left of the T-DNA, is a short region of high G C content, as evidenced by retardation of restriction-endonuclease-produced fragments on acrylamide gels and relative buoyant density.