The single transmembrane segment of gp210 is sufficient for sorting to the pore membrane domain of the nuclear envelope

The assembly of the CDS/gpCT and CDS/gpCT-20 chimeric constructs (see Fig. 1 ) was performed by similar methods with the following exceptions. The CD8 portion of the molecule was contributed by a PCR product that begins at a XbaI site in pSP65FI upstream of the CD8 eDNA and continues through CD8 to nucleotide 692. The PCR product encoding the gp210 CT and the 20-amino acid deletion both begin with nucleotide 5,485 and end with a termination codon and a BamHl linker aRer nucleotide 5,664 (CDS/gpCT) or nueleotide 5,598 (CDg/gpCT-20). The PCR product derived from the CD8 eDNA was blunt-end ligated to each of the gp210 PCR products to produce the appropriate chimeric eDNAs which were subcloned using their flanking XbaI and BamHI sites into the pSVL expression plasmid. The structure of all of the above mutants was confirmed by DNA sequencing. The CDS/EI9 chimeric eDNA within the pCMU1V plasmid (24) was kindly provided by Dr. Stefano Bonatti (Universiflt di Napoli, Napoli, Italy) with permission from Dr. Per Peterson (Scripps Clinic, La Jolla, CA).