Complete Genome Sequence Analysis of Turbot Reddish Body Iridovirus (TRBIV) and Asymptomatic Infection of TRBIV in Marine Fishes Detected by a Nested-PCR Method
|School||Ocean University of China|
|Keywords||Turbot reddish body iridovirus Genome Sequence analysis Variable region Asymptomatic infection|
Iridescent virus (iridovirus) is a class of infection the variable temperature vertebrates and invertebrates icosahedral shape, large cytoplasmic DNA virus, is important viral pathogens of aquaculture animals and wildlife. Turbot reddish body iridovirus the (turbot reddish body iridovirus, TRBIV) is a newly discovered fish iridovirus infection turbot in China and South Korea, is one of the factory farming turbot major viral pathogens. In this paper, PCR amplification, DNA cloning, such as large-scale sequencing and bioinformatics methods, measured and analyzed TRBIV complete genome sequence. The results show that the TRBIV genome is a linear double-stranded DNA, full-length to 110,104 bp, GC content of 54.99%. The whole genome containing the open reading frame of 115 possible (open reading frame, ORF), encoding polypeptide of the amino acid residues in length between 40-1168. All ORF covering 92% of the area of ??the TRBIV genome, of which 56.5% is forward coded, 43.5% for the reverse encoding. 115 ORFs in TRBIV 39 ORFs of infectious spleen and kidney necrosis virus (the the Infectious spleen and kidney necrosis virus ISKNV) Oplegnathus iridescent virus (rock bream iridovirus RBIV), brown-spotted grouper iridovirus (Orange -Spotted Grouper iridovirus, the ORF of the known functions of the OSGIV) have a high degree of similarity, and the similarity of 76-99% ,87-99% ,75-99%, respectively. These proteins can be divided into the following categories: DNA related to replication, transcription, and nucleic acid metabolism-related protein modifications related, host-related, membrane-associated. TRBIV genome dotted with many repetitive sequences, including direct repeats, inverted repeats and palindromic sequences, most of which are direct repeats and ISKNV, sea bream iridovirus (red sea bream iridovirus, RSIV), OSGIV repeating sequence is highly similar. Using TRBIV structure and function of proteins, such as the major capsid protein (major capsid protein, MCP), adenosine triphosphatase (adenosine triphosphatease, ATPase), DNA polymerase (DNA polymerase, DdDp), ribonucleotide reductase small subunit the base (ribonucleotide reductase, small chain, RNRS), the amino acid sequence of the cytosine DNA methyltransferase (Cytosine-C5-specific DNA methylase, DMet) constructed the iridovirus phylogenetic tree, and studied at the molecular level of the system evolution of TRBIV The results show that TRBIV belonging to the iridovirus Division swollen cell virus genus. Determination of the complete genome sequence in TRBIV molecular biology data and the theoretical basis for the diagnosis and the prevention and treatment of aquatic animals Iridovirus disease, opening the way for aquaculture animal iridovirus disease prevention. Will TRBIV whole genome sequences were compared with the nucleotide sequence of the the swollen cells iridescent virus genus ISKNV OSGIV, RBIV the looking TRBIV area with their differences in the sequence difference zone is swollen cells iridescent virus, within different plant types of variability between areas, that TRBIV the area of ??the most likely potential variation. Select the greater difference in five variable region, to design the reciprocal primers for PCR amplification of four different parts of different ages TRBIV at both ends of the variable region. Sequencing results show that the the different age TRBIV of four different places in five places, there are no differences. Establish of a detection TRBIV the research nested PCR method, with the method as a template for PCR amplification of different dilution factor recombinant plasmid pMD18-TRBIV ATPase, the results show the sensitivity of the nested PCR established about 10-step PCR ~ 4 times. Establish nested PCR method of our common marine aquaculture fish the TRBIV epidemiological investigation found step PCR test is negative tongue sole of the wide-body, Günther, black skirt, Africa grouper and other fish species, sets The formula length specificity 471bp DNA fragment was amplified by PCR, the fragment of the PCR product sequencing. Sequencing results in GenBank using the NCBI-Blast software search, comparison and found that the registration number as part of the fragment AY608684.1 TRBIV ATPase gene sequences with 100% homology Günther wide-body tongue sole, black The Jun Africa grouper and other four mariculture the fish from asymptomatic infection exists TRBIV, they may also TRBIV the host.