Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present s...Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.展开更多
A gene library of the infectious spleen and kidney necrosis virus(ISKNV) from mandarinfish,Siniperca chuatsi(Basilewsky) genome was established ISKNV DNA was cleaved with EcoRI,BamHI,HindⅢ,KpnI and PstI and the resul...A gene library of the infectious spleen and kidney necrosis virus(ISKNV) from mandarinfish,Siniperca chuatsi(Basilewsky) genome was established ISKNV DNA was cleaved with EcoRI,BamHI,HindⅢ,KpnI and PstI and the resulting fragments were inserted into the corresponding sites of the pBluescript Ⅱ KS plasmid vector using T4 DNA ligase Bacterial colonies harboring recombinant plasmids were selected All cloned fragments were individually identified by digestion of the recombinant plasmid DNA with different restriction enzymes and screened by hybridization of recombinant plasmid DNA to viral DNA Using these recombinant plasmids,the physical maps of the genome were constructed for BamHI,HindⅢ and KpnI restriction展开更多
文摘Viral nervous necrosis (VNN) is a worldwide disease among teleost fish. In the mainland of China, VNN was first identified in 2 species of hatchery-reared groupers, Epinephelus akaara and E. coioides. In the present study, samples were collected from larvae of E. akaara with signs of VNN in Dayawan bay which is located in southern China. The complete viral coat protein gene was amplified using extracted total RNA as template. The reverse transcription polymerase chain reaction (RT-PCR) amplification was performed using primers containing a heterologous restriction site for NotI. PCR products were cloned into pcDNA3 vector and sequenced. The results indicated that the coat protein gene of Dayawan isolate (RG-CN) was 1056 bases in length and contained a single open reading frame (ORF) of 1017 bases encoding a protein of 338 amino acids. The sequence similarities between the coat protein gene of RG-CN and other 8 isolates of NNV from Dayawan, Taiwan, Japan, Singapore and France were 79.1%-99.5% at the nucleotide level and 83.7%-100% at the amino acid level, respectively.The homology between RG-CN and the other 5 isolates from groupers was high and relatively lower between RG-CN and guppy nervous necrosis virus (GNNV), sea bass nervous necrosis virus (DIEV), but more divergences existed between RG-CN and striped jack nevous necrosis virus (SJNNV). Compared with SJNNV, RG-CN and the other 7 isolates all lacked 6 nucleotides and 2 amino acids in the same positions. Based on the result of molecular phylogenetic analysis, Dayawan isolate belongs to red-spotted grouper nervous necrosis virus (RGNNV) genotype.
文摘A gene library of the infectious spleen and kidney necrosis virus(ISKNV) from mandarinfish,Siniperca chuatsi(Basilewsky) genome was established ISKNV DNA was cleaved with EcoRI,BamHI,HindⅢ,KpnI and PstI and the resulting fragments were inserted into the corresponding sites of the pBluescript Ⅱ KS plasmid vector using T4 DNA ligase Bacterial colonies harboring recombinant plasmids were selected All cloned fragments were individually identified by digestion of the recombinant plasmid DNA with different restriction enzymes and screened by hybridization of recombinant plasmid DNA to viral DNA Using these recombinant plasmids,the physical maps of the genome were constructed for BamHI,HindⅢ and KpnI restriction
