Selection and Characterization of Packaging Cell Lines for XJ-160 Virus

Objectives: XJ-160 virus is a mosquito-derived Sindbis-like virus isolated in China. Based on its infectious clone (pBR-XJ-160) we have developed an RNA-based vector system. In this work, we constructed packaging cell lines (PCLs) for XJ-160 virus. Methods: Firstly, XJ-160 virus structural protein expression cassette pcE or pICH was constructed by cloning the gene of glycoprotein E3E26KE1 or capsid protein of XJ-160 virus into pcDNA3.1(+) or pIRES, respectively. Then the PCLs (BHK-21E+Capsid cells) for XJ-160 virus were obtained by two selections with G418 and hygromycin. Results: The results indicate that BHK-21E+Capsid cells, stably expressing E3E26KE1 protein and capsid protein of XJ-160 virus, not only highly increased packaging efficiency of the vector from XJ-160 virus, but also provided packaging function for the vector from Semliki Forest virus. Conclusion: These results suggest potential utility of the PCLs of XJ-160 virus for large-scale vector production and facilitating broad alphavirus applications. Also, the construction of PCLs for XJ-160 virus lays a basis for developing alphavirus-derived vector systems.

[1]  Cheng Shuiping,et al.  Construction and identification of reverse genetics system of Dengue type 2 virus isolated in China , 2006 .

[2]  H. Yunde.  Construction and Infection Analysis of the Full-length cDNA Clone of XJ-160 Virus,the First Sindbis Virus Isolated in China , 2005 .

[3]  P. D. Felipe.  Skipping the co-expression problem: the new 2A "CHYSEL" technology. , 2004 .

[4]  P. de Felipe.  Skipping the co-expression problem: the new 2A "CHYSEL" technology , 2004, Genetic vaccines and therapy.

[5]  L. Guo-dong,et al.  Construction of Replicon Expression Vector Derived from XJ-160 Virus , 2003 .

[6]  L. Li,et al.  Isolation and complete nucleotide sequence of a Chinese Sindbis-like virus. , 2000, The Journal of general virology.

[7]  T. Dubensky,et al.  Stable alphavirus packaging cell lines for Sindbis virus and Semliki Forest virus-derived vectors. , 1999, Proceedings of the National Academy of Sciences of the United States of America.

[8]  R. Johnston,et al.  Replicon-helper systems from attenuated Venezuelan equine encephalitis virus: expression of heterologous genes in vitro and immunization against heterologous pathogens in vivo. , 1997, Virology.

[9]  E. Frolova,et al.  Packaging signals in alphaviruses , 1997, Journal of virology.

[10]  C. Rice,et al.  Alphavirus-based expression vectors: strategies and applications. , 1996, Proceedings of the National Academy of Sciences of the United States of America.

[11]  H. Huang,et al.  Sindbis virus vectors for expression in animal cells. , 1996, Current opinion in biotechnology.

[12]  C. Rice,et al.  Sindbis virus expression vectors: packaging of RNA replicons by using defective helper RNAs , 1993, Journal of virology.

[13]  P. Liljeström,et al.  A New Generation of Animal Cell Expression Vectors Based on the Semliki Forest Virus Replicon , 1991, Bio/Technology.

[14]  T. Gadek,et al.  Evidence for specificity in the encapsidation of Sindbis virus RNAs , 1989, Journal of virology.

[15]  C. Rice,et al.  Sindbis virus: an efficient, broad host range vector for gene expression in animal cells. , 1989, Science.

[16]  S. Goff,et al.  A safe packaging line for gene transfer: separating viral genes on two different plasmids , 1988, Journal of virology.

[17]  M. Nicolson,et al.  Replication-defective chimeric helper proviruses and factors affecting generation of competent virus: expression of Moloney murine leukemia virus structural genes via the metallothionein promoter , 1987, Molecular and cellular biology.