The complete nucleotide sequence of two human T-cell leukaemia type III (HTLV-III) proviral DNAs each have four long open reading frames, the first two corresponding to the gag and pol genes. The fourth open reading frame encodes two functional polypeptides, a large precursor of the major envelope glycoprotein and a smaller protein derived from the 3'-terminus long open reading frame analogous to the long open reading frame (lor) product of HTLV-I and -II.

A DNA sequencing system based on the use of a novel set of four chain-terminating dideoxynucleotides, each carrying a different chemically tuned succinylfluorescein dye distinguished by its fluorescent emission is described. Avian myeloblastosis virus reverse transcriptase is used in a modified dideoxy DNA sequencing protocol to produce a complete set of fluorescence-tagged fragments in one reaction mixture. These DNA fragments are resolved by polyacrylamide gel electrophoresis in one sequencing lane and are identified by a fluorescence detection system specifically matched to the emission characteristics of this dye set. A scanning system allows multiple samples to be run simultaneously and computer-based automatic base sequence identifications to be made. The sequence analysis of M13 phage DNA made with this system is described.

No abstract

We describe experiments using the swine herpesvirus, pseudorabies virus (PRV), as a vector for expression of hybrid membrane protein genes. In particular, we present the construction and analysis of three infectious PRV mutants expressing chimeric viral membrane proteins composed of portions of the PRV envelope glycoprotein gIll and of the human retrovirus, human immunodeficiency virus type 1 (HIV-1), envelope glycoproteins gpl20 and gp4l. All of the chimeric genes contain the transcription control sequences and the first 157 codons of PRV gIII (known to contain signals sufficient for efficient export of the encoded peptide out of the cell) fused to different regions of the HIV-1 envelope. The mutant viruses express novel glycosylated fusion proteins that are immunoprecipitated by polyvalent sera specific for gIII, as well as acquired immunodeficiency syndrome patient sera. The levels of expression are lower than expected due primarily to instability or altered processing of the hybrid mRNA. We could not detect cleavage of chimeric proteins carrying the gp120-gp41 protease processing site. The use of localization signals contained within herpesvirus membrane proteins to direct chimeric proteins to desired cellular locations is discussed.