Genomic localization of the gene encoding a 32-kDa capsid protein of human cytomegalovirus

Abstract

We have determined the map position of a viral gene encoding a 32-kDa late structural protein of human cytomegalovirus (HCMV) using a murine monoclonal antibody. This monoclonal antibody was reactive with two protein bands of 32 and 27 kDa in HCMV-infected cell lysates and with a single 32-kDa protein band in HCMV virions as detected by immunoblot analysis. When purified HCMV envelope preparation was used for immunoblotting, the monoclonal antibody did not display a detectable band. We used this monoclonal antibody to screen a cDNA library that was constructed from poly(A)+ RNA of late HCMV-infected cells and cloned into the expression vector lambda gt11. A cDNA clone that expressed an immunoreactive epitope of the late HCMV protein fused to beta-galactosidase was identified. Probing the restriction digests of HCMV (Towne and AD169) DNA with insert DNA from the immunoreactive lambda gt11 clone permitted us to localize the coding sequence within the long unique region between map coordinates of 0.62 and 0.64 of HCMV Towne and AD169 genomes. Using the same probe, a single transcript of 1.4 kb was detected in total RNA from HCMV-infected cells at late times after infection.