Abstract
We employed the staggered extension process (StEP) to shuffle the S1 genes from four infectious bronchitis virus (IBV) strains representing four unique serotypes. Upon creating a shuffled S1 gene library, we randomly selected 25 clones and analyzed them by DNA sequencing. In total, eleven clones contained novel S1 gene recombinants. Based on sequence data, each recombinant was unique and contained a full-length open reading frame. The average number of crossovers per recombinant was 5 and the average number of point mutations was 1.3, leading mostly to non-synonymous amino acid changes. No recombinant contained sequences from all four parental genes and no recombinant contained any sequence from the distantly related Delaware 072 strain. Our data suggests that recombination between distantly related IBV strains within the S1 gene probably does not readily occur. This finding is extremely important in light of the common industry vaccination practice of mixing different live-attenuated IBV strains.
Keywords: DNA shuffling, infectious bronchitis virus (IBV), staggered extension process (StEP), S1 gene
References
- 1.King D.J. and Cavanagh D., in Calnek B.W. (ed.), Diseases of Poultry, Iowa State University Press, Ames, 1991, pp. 471–484.
- 2.Wang L., Junker D., Collisson E.W. Virology. 1993;192(2):710–716. doi: 10.1006/viro.1993.1093. [DOI] [PubMed] [Google Scholar]
- 3.Jia W., Karaca K., Parrish C.R., Naqi S.A. Arch Virol. 1995;140(2):259–271. doi: 10.1007/BF01309861. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 4.Kottier S.A., Cavanagh D., Britton P. Virology. 1995;213(2):569–580. doi: 10.1006/viro.1995.0029. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 5.Lee C.W., Jackwood M.W. Arch Virol. 2000;145(10):2135–2148. doi: 10.1007/s007050070044. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 6.Lee C.W., Jackwood M.W. Virus Res. 2001;80(1–2):33–39. doi: 10.1016/S0168-1702(01)00345-8. [DOI] [PubMed] [Google Scholar]
- 7.Lee C.W., Jackwood M.W. Virus Genes. 2001;22(1):85–91. doi: 10.1023/A:1008138520451. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 8.Stemmer W.P. Proc Natl Acad Sci USA. 1994;91(22):10747–10751. doi: 10.1073/pnas.91.22.10747. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 9.Stemmer W.P. Nature. 1994;370(6488):389–391. doi: 10.1038/370389a0. [DOI] [PubMed] [Google Scholar]
- 10.Soong N.W., Nomura L., Pekrun K., Reed M., Sheppard L., Dawes G., Stemmer W.P. Nat Genet. 2000;25(4):436–439. doi: 10.1038/78132. [DOI] [PubMed] [Google Scholar]
- 11.Miyazaki K. Nucleic Acids Res. 2002;30(24):e139, 2002. doi: 10.1093/nar/gnf139. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 12.Zhao H., Giver L., Shao Z., Affholter J.A., Arnold F.H. Nat Biotechnol. 1998;16(3):258–261. doi: 10.1038/nbt0398-258. [DOI] [PubMed] [Google Scholar]
- 13.Jungherr, in Proc 60th Ann Mtg U.S.LS.A, 1956
- 14.Gelb J., Jr, Perkins B.E., Rosenberger J.K., Allen P.H. Avian Dis. 1981;25(3):655–666. [PubMed] [Google Scholar]
- 15.Gelb J., Jr, Keeler C.L., Jr, Nix W.A., Rosenberger J.K., Cloud S.S. Avian Dis. 1997;41(3):661–669. [PubMed] [Google Scholar]
- 16.Senne D.A., in Swayne, D.E. (ed.), A Laboratory Manual for the Isolation and Identification of Avian Pathogens, American Association of Avian Pathologists: Kennett Square, 1998, pp. 235–240
- 17.Kwon H.M., Jackwood M.W., Gelb J., Jr Avian Dis. 1993;37(1):194–202. [PubMed] [Google Scholar]
- 18.Lee C.W., Hilt D.A., Jackwood M.W. Avian Dis. 2000;44(3):650–654. [PubMed] [Google Scholar]
- 19.Niesters H.G., Bleumink-Pluym N.M., Osterhaus A.D., Horzinek M.C., van der Zeijst B.A. Virology. 1987;161(2):511–519. doi: 10.1016/0042-6822(87)90145-0. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 20.Cavanagh D., Davis P.J., Mockett A.P. Virus Res. 1988;11(2):141–150. doi: 10.1016/0168-1702(88)90039-1. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 21.Moore K.M., Jackwood M.W., Hilt D.A. Arch Virol. 1997;142(11):2249–2256. doi: 10.1007/s007050050239. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 22.Moore K.M., Bennett J.D., Seal B.S., Jackwood M.W. Virus Genes. 1998;17(1):63–83. doi: 10.1023/A:1008057118625. [DOI] [PMC free article] [PubMed] [Google Scholar]
- 23.Jackwood M.W., Hilt D.A., Callison S.A., Lee C.W., Plaza H., Wade E. Avian Dis. 2001;45(2):366–372. [PubMed] [Google Scholar]
- 24.Chou P.Y., Fasman G.D. Adv Enzymol Relat Areas Mol Biol. 1978;47:45–148. doi: 10.1002/9780470122921.ch2. [DOI] [PubMed] [Google Scholar]
- 25.Kant A., Koch G., van Roozelaar D.J., Kusters J.G., Poelwijk F.A., van der Zeijst B.A. J Gen Virol. 1992;73(Pt3):591–596. doi: 10.1099/0022-1317-73-3-591. [DOI] [PubMed] [Google Scholar]