Expression of a recombinant DNA gene coding for the vesicular stomatitis virus nucleocapsid protein

A cDNA clone containing the entire vesicular stomatitis virus nucleocapsid gene was assembled by fusing portions of two partial clones. When the cDNA clone was inserted into a new general-purpose eucaryotic expression vector and introduced into appropriate host cells, abundant N-protein synthesis ensued. The expressed protein was indistinguishable from authentic N protein produced during vesicular stomatitis virus infections. The recombinant N protein was recognized by a polyclonal antibody and two different monoclonal antibodies and could not be resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis from authentic N. Our results suggest that the recombinant N protein produced in transfected cells rapidly aggregates into high-molecular-weight complexes in the absence of vesicular stomatitis virus genomic RNA.

[1]  T. Maniatis,et al.  Identification of DNA sequences required for transcription of the human α1-globin gene in a new SV40 host-vector system , 1981, Cell.

[2]  J. Keene,et al.  The origins of defective interfering particles of the negative-strand RNA viruses , 1981, Cell.

[3]  M. Botchan,et al.  Inhibition of SV40 replication in simian cells by specific pBR322 DNA sequences , 1981, Nature.

[4]  R. Wu,et al.  An improved procedure for utilizing terminal transferase to add homopolymers to the 3' termini of DNA. , 1981, Nucleic acids research.

[5]  J. Greene,et al.  Nucleotide sequences of the mRNA's encoding the vesicular stomatitis virus N and NS proteins , 1981, Journal of virology.

[6]  M. Leppert,et al.  Interaction of VSV leader RNA and nucleocapsid protein may control VSV genome replication , 1981, Cell.

[7]  Y. Gluzman SV40-transformed simian cells support the replication of early SV40 mutants , 1981, Cell.

[8]  D. Nayak,et al.  Defective interfering influenza viruses and host cells: establishment and maintenance of persistent influenza virus infection in MDBK and HeLa cells , 1980, Journal of virology.

[9]  Stanley N Cohen,et al.  3′-End labeling of DNA with [α-32P] cordycepin-5′-triphosphate , 1980 .

[10]  George R. Stark,et al.  Regulation of Simian Virus 40 Transcription: Sensitive Analysis of the RNA Species Present Early in Infections by Virus or Viral DNA , 1979, Journal of virology.

[11]  G. Ryffel,et al.  Cloning and characterization of synthetic sequences from the Xenopus iaevis vitellogenin structural gene. , 1978, Developmental biology.

[12]  R. Colonno,et al.  Complete nucleotide sequence of the leader rna synthesized in vitro by vesicular stomatitis virus , 1978, Cell.

[13]  J. Keene,et al.  Nucleotide sequence homology at the 3' termini of RNA from vesicular stomatitis virus and its defective interfering particles. , 1978, Proceedings of the National Academy of Sciences of the United States of America.

[14]  R T Schimke,et al.  Synthesis of full length cDNAs from four partially purified oviduct mRNAs. , 1978, The Journal of biological chemistry.

[15]  S. Emerson,et al.  In vitro synthesis of a unique RNA species by a T particle of vesicular stomatitis virus , 1977, Journal of virology.

[16]  R. Lazzarini,et al.  The relationship between autointerference and the replication of defective interfering particle. , 1977, Virology.

[17]  R. Colonno,et al.  A unique RNA species involved in initiation of vesicular stomatitis virus RNA transcription in vitro , 1976, Cell.

[18]  D. Hogness,et al.  Colony hybridization: a method for the isolation of cloned DNAs that contain a specific gene. , 1975, Proceedings of the National Academy of Sciences of the United States of America.

[19]  R. Lazzarini,et al.  Covalently linked message and anti-message (genomic) RNA from a defective vesicular stomatitis virus particle. , 1975, Journal of molecular biology.

[20]  F. Brown,et al.  Model for Vesicular Stomatitis Virus , 1972, Journal of virology.

[21]  K. Hummeler,et al.  Morphogenesis of the Nucleoprotein of Vesicular Stomatitis Virus , 1970, Journal of virology.

[22]  W. Gilbert,et al.  Sequencing end-labeled DNA with base-specific chemical cleavages. , 1980, Methods in enzymology.

[23]  D. Botstein,et al.  Advanced bacterial genetics , 1980 .

[24]  R. Kolter,et al.  Plasmid cloning vehicles derived from plasmids ColE1, F, R6K, and RK2. , 1979, Methods in enzymology.