Skip to main content
Elsevier - PMC COVID-19 Collection logoLink to Elsevier - PMC COVID-19 Collection
. 2004 Feb 6;151(1):41–49. doi: 10.1016/0042-6822(86)90102-9

Sequence analysis of the porcine transmissible gastroenteritis coronavirus nucleocapsid protein gene

Paul A Kapke 1,1, David A Brian 1,2
PMCID: PMC7131278  PMID: 3008432

Abstract

The 3′ end of the 20-kb genome of the Purdue strain of porcine transmissible gastroenteritis coronavirus (TGEV) was copied into eDNA after priming with oligo(dT) and the double-stranded product was cloned into the PstI site of the pUC9 vector. One clone of 2.0-kb contained part of the poly(A) tail and was sequenced in its entirety using the chemical method of Maxam and Gilbert. Another clone of 0.7 kb also contained part of the poly(A) tail and was sequenced in part to confirm the primary structure of the most 3′ end of the genome. Two potential, nonoverlapping genes were identified within the 3′-terminal 1663-base sequence from an examination of open reading frames. The first gene encodes a 382-amino acid protein of 43,426 mol wt, that is the apparent nucleocapsid protein on the basis of size, chemical properties, and amino acid sequence homology with other coronavirus nucleocapsid proteins. It is flanked on its 5′ side by at least part of the matrix protein gene. The second encodes a hypothetical 78-amino acid protein of 9101 mol wt that is hydrophobic at both ends. A 3′-proximal noncoding sequence of 276 bases was also determined and a conserved stretch of 9 nucleotides near the poly(A) tail was found to be common among TGEV, the mouse hepatitis coronavirus, and the avian infectious bronchitis coronavirus.

References

  1. Armstrong J., Niemann H., Smeekens S., Rottier P., Warren G. Sequence and topology of a model intracellular membrane protein, E1 glycoprotein, from a coronavirus. Nature (London) 1984;308:751–752. doi: 10.1038/308751a0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Armstrong J., Smeekens S., Rottier P. Sequence of the nucleocapsid gene from murine coronavirus MHV-A59. Nucleic Acids Res. 1983;11:833–891. doi: 10.1093/nar/11.3.883. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Boursnell M.E.G., Binns M.M., Foulds I.J., Brown T.D.K. Sequences of the nucleocapsid genes from two strains of avian infectious bronchitis virus. J. Gen. Virol. 1985;66:573–580. doi: 10.1099/0022-1317-66-3-573. [DOI] [PubMed] [Google Scholar]
  4. Boursnell M.E.G., Brown T.D.K. Sequencing of coronavirus IBV genomic RNA: A 195-base open reading frame encoded by mRNA B. Gene. 1984;29:87–92. doi: 10.1016/0378-1119(84)90169-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Brian D.A., Dennis D.E., Guy J.S. Genome of porcine transmissible gastroenteritis virus. J. Virol. 1980;34:410–415. doi: 10.1128/jvi.34.2.410-415.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Brown T.D.K., Boursnell M.E.G. Avian infectious bronchitis virus genomic RNA contains sequence homologies at the intergenic boundaries. Virus Res. 1984;1:15–24. [Google Scholar]
  7. Budzilowicz C.J., Wilczynski S.P., Weiss S.R. Three intergenic regions of coronavirus mouse hepatitis virus strain A59 genome RNA contain a common nucleotide sequence that is homologous to the 3′ end of the viral mRNA leader sequence. J. Virol. 1985;53:834–840. doi: 10.1128/jvi.53.3.834-840.1985. [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Dennis D.E., brian D.A. RNA-dependent RNA polymerase activity in coronavirus-infected cells. J. Virol. 1982;42:153–164. doi: 10.1128/jvi.42.1.153-164.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Garwes D.J., Pocock D.H. The polypeptide structure of transmissible gastroenteritis virus. J. Gen. Virol. 1975;29:25–34. doi: 10.1099/0022-1317-29-1-25. [DOI] [PubMed] [Google Scholar]
  10. Gubler U., Hoffman B.J. A simple and very efficient method for generating cDNA libraries. Gene. 1983;25:263–269. doi: 10.1016/0378-1119(83)90230-5. [DOI] [PubMed] [Google Scholar]
  11. Hanahan D. Studies on transformation of Escherichia coli with plasmids. J. Mol. Biol. 1983;166:557–580. doi: 10.1016/s0022-2836(83)80284-8. [DOI] [PubMed] [Google Scholar]
  12. Heidecker G., Messing J. Sequence analysis of zein cDNAs obtained by an efficient mRNA method. Nucleic Acids Res. 1983;11:4891–4906. doi: 10.1093/nar/11.14.4891. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Hu S., Bruszewski J., Boom T., Souza L. Cloning and expression of the surface glycoprotein gp195 of porcine transmissible gastroenteritis virus. In: Chanock R.M., Lerner R.A., editors. Modern Approaches to Vaccines. Cold Spring Harbor Laboratory; Cold Spring Harbor, N.Y: 1984. pp. 219–223. [Google Scholar]
  14. Kado C.I., Liu S.T. Rapid procedure for detection and isolation of large and small plasmids. J. Bacteriol. 1981;145:1365–1373. doi: 10.1128/jb.145.3.1365-1373.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Kozak M. Comparison of initiation of protein synthesis in procaryotes, eucaryotes, and organelles. Microbiol. Rev. 1983;47:1–45. doi: 10.1128/mr.47.1.1-45.1983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lai M.M.C., Brayton P.R., Armen R.C., Patton C.D., Stohlman S.A. Mouse hepatitis virus A59 messenger RNA structure and genetic localization of the sequence divergence from the hepatotropic strain MHV 3. J. Virol. 1981;39:823–834. doi: 10.1128/jvi.39.3.823-834.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Leibowitz J.L., Weiss S.R., Paavola E., Bond C.W. Cell-free translation of murine coronavirus RNA. J. Virol. 1982;43:905–913. doi: 10.1128/jvi.43.3.905-913.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Maniatis T., Fritsch E.F., Sambrook J. Cold Spring Harbor Laboratory; Cold Spring Harbor, N.Y: 1982. (Molecular Cloning: A Laboratory Manual). [Google Scholar]
  19. Maxam A.M., Gilbert W. Sequencing end-labeled DNA with base-specific chemical cleavages. In: Grossman L., Moldave K., editors. Vol. 65. Academic Press; Orlando, Fla: 1980. pp. 499–560. (Methods in Enzymology). [DOI] [PubMed] [Google Scholar]
  20. Peacock S.L., McIver C.M., Monohan J.J. Transformation of E. coli using homopolymer-linked plasmid chimeras. Biochim. Biophys. Acta. 1981;655:243–250. doi: 10.1016/0005-2787(81)90014-9. [DOI] [PubMed] [Google Scholar]
  21. Pedersen N.C., Ward J., Mengeling W.L. Antigenic relationship of the feline infections peritonitis virus to coronaviruses of other species. Arch. Virol. 1978;58:45–53. doi: 10.1007/BF01315534. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Queen C., Korn L.J. A comprehensive sequence analysis program for the IBM personal computer. Nucleic Acids Res. 1984;12:581–599. doi: 10.1093/nar/12.1part2.581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Rottier P.J.M., Horzinek M.C., van der Zelist B.A.M. Translation of three mouse hepatitis virus (MHV-A59) subgenomic RNAs in Xenopus laevis oocytes. J. Virol. 1981;38:20–26. doi: 10.1128/jvi.38.1.20-26.1981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Roychoudhury R., Wu R. Terminal transferase-catalyzed addition of nucleotides to the 3′ termini of DNA. In: Grossman L., Moldave K., editors. Vol. 65. Academic Press; Orlando, Fla: 1980. pp. 43–62. (Methods of Enzymology). [DOI] [PubMed] [Google Scholar]
  25. Skinner M.A., Ebner D., Siddell S.G. Coronavirus MHV-JHM mRNA 5 has a sequence arrangement which potentially allows translation of a second, downstream open reading frame. J. Gen. Virol. 1985;66:581–592. doi: 10.1099/0022-1317-66-3-581. [DOI] [PubMed] [Google Scholar]
  26. Skinner M.A., Siddell S.G. Coronavirus JHM: Nucleotide sequence of the mRNA that encodes nucleocapsid protein. Nucleic Acids Res. 1983;11:5045–5054. doi: 10.1093/nar/11.15.5045. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Smith H.O., Birnstiel M.L. A simple method for DNA restriction site mapping. Nucleic Acids Res. 1976;3:2387–2398. doi: 10.1093/nar/3.9.2387. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Stern D.F., Kennedy S.I.T. Coronavirus multiplication strategy. II. Mapping the avian infectious bronchitis virus intracellular RNA species to the genome. J. Virol. 1980;36:440–449. doi: 10.1128/jvi.36.2.440-449.1980. [DOI] [PMC free article] [PubMed] [Google Scholar]
  29. Stohlman S.A., Lai M.M.C. Phosphoproteins of murine hepatitis virus. J. Virol. 1979;32:672–675. doi: 10.1128/jvi.32.2.672-675.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Virology are provided here courtesy of Elsevier

RESOURCES