On the basis of the sequence and analysis of genome from the orange-spotted nervous necrosis virus（ OGNNV）, China strain, a pair of special primers were designed according to the nucleotide sequences of RNA2 from OG...On the basis of the sequence and analysis of genome from the orange-spotted nervous necrosis virus（ OGNNV）, China strain, a pair of special primers were designed according to the nucleotide sequences of RNA2 from OGNNV. The major capsid protein （ MCP）gene of OGNNV was cloned by means of reverse-transcriptase polymerase chain reaction （RT-PCR） and ligated into the pET32a expression plasmid. The MCP gene of OGNNV was 1 017 bases, encoded a protein of 338 amino acid with a molecular mass of 37.1 kDa. Recombinant protein with a molecular mass of 57.4 kDa was expressed in E. coli BL21 （DE3）. Vaccine was prepared from the recombinant protein expressed in recombinant cells. The juvenile orange-spotted groupers （8 cm in average length） were immunized by intraperitoneal injection. Group A was challenged with infected tissue filtrates 25 d post-vaccination. The mortality in the vaccined group （ A1,30% ） was a little higher than the unvaccined group （ B2, 27.8% ）. Group B was challenged after three vaccine injections. The mortality in the vaccined group （B1, 16.7% ） was lower than the unvaccined group （132, 27.8% ）, And the relative percentage survival （RPS） value of vaccined group, compared with the unvaccined group, was 40%. The anti-recombinant protein sera with a 1 ： 100 dilution were mixed with double volume of infected tissue filtrates and incubated at 4 ℃ for 12 h and then intramuscularly injected into the juvenile orange-spotted grouper. Treatment of infected tissue filtrates with anti-recombinant protein serum resulted in a significantly lower mortality of fish （ Group C1, mortality of 18.18% ）, compared with the fish （ Group C2, mortality of 40% ） which received infected tissue filtrates treated with control serum. Results implied the potential use of the capsid protein in immunization against OGNNV.展开更多
基金This work was supported by the National"863"Projects of China under contract No.2001AA601010the Natural Science Foundation of Jinan University,China under contract No.51204062.
文摘On the basis of the sequence and analysis of genome from the orange-spotted nervous necrosis virus（ OGNNV）, China strain, a pair of special primers were designed according to the nucleotide sequences of RNA2 from OGNNV. The major capsid protein （ MCP）gene of OGNNV was cloned by means of reverse-transcriptase polymerase chain reaction （RT-PCR） and ligated into the pET32a expression plasmid. The MCP gene of OGNNV was 1 017 bases, encoded a protein of 338 amino acid with a molecular mass of 37.1 kDa. Recombinant protein with a molecular mass of 57.4 kDa was expressed in E. coli BL21 （DE3）. Vaccine was prepared from the recombinant protein expressed in recombinant cells. The juvenile orange-spotted groupers （8 cm in average length） were immunized by intraperitoneal injection. Group A was challenged with infected tissue filtrates 25 d post-vaccination. The mortality in the vaccined group （ A1,30% ） was a little higher than the unvaccined group （ B2, 27.8% ）. Group B was challenged after three vaccine injections. The mortality in the vaccined group （B1, 16.7% ） was lower than the unvaccined group （132, 27.8% ）, And the relative percentage survival （RPS） value of vaccined group, compared with the unvaccined group, was 40%. The anti-recombinant protein sera with a 1 ： 100 dilution were mixed with double volume of infected tissue filtrates and incubated at 4 ℃ for 12 h and then intramuscularly injected into the juvenile orange-spotted grouper. Treatment of infected tissue filtrates with anti-recombinant protein serum resulted in a significantly lower mortality of fish （ Group C1, mortality of 18.18% ）, compared with the fish （ Group C2, mortality of 40% ） which received infected tissue filtrates treated with control serum. Results implied the potential use of the capsid protein in immunization against OGNNV.
