Abstract: In order to obtain and analyze the complete genome sequences of enzootic nasal tumor virus 2 (ENTV-2) in goat intranasal tumor tissues obtained from a Fujian farm, six specific fragments of the ENTV-2 were amplified from these tumor tissues by RT-PCR. The products were then sequenced and assembled using biological software DNAStar to obtain the whole genome sequences. We found that whole length of the viral genome was 7 443 bp, and the virus was named as ENTV-2-FJ. The structure of ENTV-2-FJ genome is similar to other retroviruses, which has a 5'-U5-gag-pro-pol-env-U3-3' canonical structure with four open reading frames, flanked by non-coding regions and terminal repeats sequence. In order to prepare rabbit anti-ENTV-2 gag protein polyclonal antibody, a pair of specific primers was designed and used to clone the gag gene that was then ligated to pET-21a (+) vector. The recombinant plasmid pET-21a-gag was constructed, transferred into BL21 (DE3) competent cells, and the recombinant fusion protein with a molecular weight of about 70 ku was successfully induced by IPTG. Western blot analysis showed that the prepared rabbit anti-gag protein antibody specifically detected the gag protein in the tumor tissues in diseased goats and cells expressing gag gene. Furthermore, we generated K562 cell lines overexpressing the gag gene and conducted tumorigenic experiments on nude mice. The results showed that overexpression of gag gene promoted tumor growth through activation of JAK2-STAT5 signaling pathway. Taken together, this study provides the complete genomic sequences of ENTV-2-FJ and rabbit polyclonal antibodies against viral gag protein, and reveals an important role of the gag protein in tumorigenesis mediated by ENTV-2-FJ.

Key words: ENTV-2, whole gene sequence, polyclonal antibody preparation, gag, pathogenicity research