Chemistry: molecular biology and microbiology – Spore forming or isolating process
Patent
1990-02-01
1995-03-07
Weimar, Elizabeth C.
Chemistry: molecular biology and microbiology
Spore forming or isolating process
4353201, 530395, 536 2372, C12N 500, C12N 1500, C07K 1300, C07H 1700
Patent
active
053957598
DESCRIPTION:
BRIEF SUMMARY
This invention relates to rotavirus, genes, gene segments, cloned genes and segments and products obtained therefrom including diagnostic reagents and vaccines.
Rotavirus is now recognized by the World Health Organization as a major cause of infantile gastroenteritis, and a high priority has been placed on control of this disease by the production of a suitable vaccine (1). Cross-neutralization tests indicate four (or possibly five) (2-4) serotypes of human rotavirus and animal studies appear to show little cross-protection between serotypes (5). Thus a potential vaccine may have to incorporate all the known human serotypes. The virus serotype has recently been shown to be determined by the major outer shell glycoprotein (6-10) (a virus surface protein), and the gene segments coding for this protein from a bovine (UK) and a simian (SA11) rotavirus have recently been sequenced (11, 12). To date however, no such gene from human rotavirus has been analysed. We therefor cloned and sequenced the gene encoding this protein from a human rotavirus. Hu/5 (isolated in Melbourne, Australia) belonging to serotype 2.
The present invention provides a human rotavirus gene and a cloned human rotavirus gene, the use of such genes to obtain expression of antigenic viral proteins such as in bacterial/procaryotic or eucaryotic expression systems and the expression products obtained and further including vaccines and diagnostic reagents obtained therefrom.
The present invention also provides the dsRNA gene segment coding for the major outer capsid glycoprotein of a human rotavirus and, without prejudice to the generality of the foregoing, that human rotavirus may be Hu/Australia/5/77 (serotype 2), a DNA copy of same, a clone thereof, or a vector or a host cell containing same, peptide sequences obtained therefrom. Of particular interest are vectors such as plasmids obtained therefrom and host cell containing same.
The present invention also provides a material comprising a nucleotide sequence coding for at least part of the major outer capsid glycoprotein of a rotavirus.
In one instance the present invention provides at least one of the nucleotide sequences from nucleotide numbers 291-357, 480-513 and 657-720 of a rotavirus major outer capsid glycoprotein gene.
In another instance the present invention provides at least one of the amino acid sequences from amino acid numbers 82-103, 144-155 and 204-224 for which the nucleotide sequences of a rotavirus major outer capsid glycoprotein gene code.
In a particularly preferred instance the present invention provides a material comprising a nucleotide sequence encoding, or an amino acid sequence being,
a. an amino acid sequence of 22 amino acids commencing CLYYP and terminating TLS,
b. an amino acid sequence of 12 amino acids commencing YD and terminating SEL, or
c. an amino acid sequence of 21 amino acids commencing GIGC and terminating EKL,
and derived from a nucleotide sequence coding for a major outer capsid glycoprotein of a rotavirus.
Specific portions of cloned genes are provided by this invention and the invention extends to products obtained therefrom including anti-sera or anti-bodies prepared by utilization of such amino acid sequences.
This invention will be exemplified by the following description.
MATERIALS AND METHODS
Virus growth and purification
The human rotavirus Hu/5 (Hu/Australia/5/77) (13) was grown in MA104 cells and purified as described previously (14).
Cloning rotavirus cDNA
The procedure for producing cDNA from rotavirus dsRNA, and cloning it into the PstI site of the plasmid pBR322 has been described previously by Dyall-Smith et al. (15).
Identification of cloned copies of the major outer shell glycoprotein gene
of Hu/5 rotavirus
Since the UK bovine rotavirus gene encoding the major outer shell glycoprotein (gene 8 of this virus) had previously been cloned (11), this was used to screen the Hu/5 clones. To eliminte pBR322 sequences, the UK gene 8 clone was digested with PstI and the insert separated by agarose gel electrophoresis. The insert was
REFERENCES:
Green, Kim Y. et al., (1987) "Comparison of the Amino Acid Sequences of the Major Neutralization Protein of Four Human Rotavirus Serotypes", Virology 161, 153-159.
Both et al. (1983) Serotype Specific Glycoprotein of Simian 11 Rotavirus: Coding Assignment and Gene Sequence, PNAS 80, 3091-3095.
Estes et al. (1989) "Rotavirus Gene Structure and Function" Micro-Biological Reviews 53, 410-449.
Thouless, M. E., et al. (1977) Serological relationships between rotaviruses from different species as studies by complement fixation and neutralization. Arch. Virol. 53: 287-294.
Dyall-Smith, M. L., et al. (1981) Gene-coding assignments of rotavirus double-stranded RNA segments 10 and 11. J. Virol. 38: 1099-1103.
Kalica, A. R., et al., (1981) Genes of human (strain Wa) and bovine (strain UK) rotaviruses that code for neutralization and subgroup antigens. Virology 112: 385-390.
Wyatt, R. G., et al. (1982) Definition of human rotavirus serotypes by plaque reduction assay. Infect. Immun. 37:110-115.
Sonza, S., et al., (1983) Derivation of neutralizing monoclonal antibodies against rotavirus. J. Virol. 45:1143-1146.
Dyall-Smith, M. L., et al. (1983) Gene mapping of rotavirus double-stranded RNA segments by northern blot hybridization: Application to segments 7, 8, and 9. J. Virol. 46:317-320.
Dyall-Smith, M. L., et al., (1983) Cloning and sequencing of UK bovine rotavirus gene segment 7: marked sequence homology with simian rotavirus gene segment 8. Nucelic Acids Res. 11:3351-3362.
Elleman, T. C., et al., (1983) Nucleotide sequence of the gene encoding the serotype-specific glycoprotein of UK bovine rotavirus Nucleic Acids Res. 11: 4689-4701.
Dyall-Smith Michael L.
Holmes Ian H.
Crouch Deborah
The University of Melbourne
Weimar Elizabeth C.
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