Using real-time quantitative polymerase chain reaction as a platform for the analysis of gene expression of orange spotted grouper infected with iridovirus
Date Issued
2011
Date
2011
Author(s)
Liu, Yu-Cheng
Abstract
Grouper iridovirus (GIV), a double-strand DNA virus, can infect with all life stages of orange-spotted grouper (Epinephelus coioides), and causes groupers serious death. It mainly attacks immune system of host, such as spleen and kidney. Until now, the total 140 kb genome size of GIV had been sequenced and the research of several virus genes expression upon infection had been studied. However, investigation of host gene expression during GIV infection is still know very little. The grouper gene expression profiles after different kinds of challenges including GIV, lipopolysaccharide, and poly I:C using GIV-infected grouper kidney cDNA microarray had been established in our lab previously.
We reanalyzed the previous array data of GIV-challenged only, and the newly 39 up-regulated and 48 down-regulated genes under two-fold thresholud were selected. To validate the result of microarray analysis, the quantitative real-time RT-PCR was performed. The kidney parts of non-injected control groupers and GIV-, lipopolysaccharide-, poly I:C-treated of groupers were collected at 1, 3, 5 day post-injection and the total RNAs were extracted for further RT-qPCR. Twenty four candidate genes from microarray analysis and another 6 immune-related genes were estimated. Fifteen genes including RNA helicase DHX58 homolog, CD9 antigen, interferon stimulated gene 15, HECT E3 ubiquitin ligase, reverse transcriptase-like protein, vig-1, very large inducible GTPase-1, urokinase plasminogen activator surface receptor, 26S proteasome non-ATPase regulatory subunit 1 homolog, interleukin-1β, interleukin-8, interferon-induced GTP-binding protein MxI, tumor-necrosis factor α and cyclooxygenase-2 are significant expression up to 20 fold compared to non-injected control candidate gene, but UDP-glucose 6-dehydrogenase is suppressed down to around 100 fold after GIV-injected treatment. Based on this experiment, we believe that those genes with significant expressed or suppressed may participate in the host-viral interaction.
Subjects
grouper
qpcr
giv
Type
thesis
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