Abortive Infection of Snakehead Fish Vesiculovirus in ZF4 Cells Was Associated with the RLRs Pathway Activation by Viral Replicative Intermediates
Abstract
:1. Introduction
2. Results
2.1. Characterization of DrLGP2a and DrLGP2b cDNAs
2.2. The mRNA and cRNA Expressions of the Viral N and G Genes in ZF4 Cells Infected with SHVV
2.3. Temporal Expression of IFN and Mx in ZF4 after Infection with SHVV
2.4. Temporal mRNA Expressions of RIG-I, MDA5, DrLGP2T and DrLGP2a in ZF4 Cells after Infection with SHVV
2.5. Temporal Expressions of DrLGP2a and DrLGP2b Proteins in ZF4 after the Infection with SHVV
Names | Sequence (5'→3') | Application |
---|---|---|
LGP2-FW | CATTAATGAGAAGGGCATCGTT | DrLGP2 core fragment |
LGP2-BW | GGCTCAGATTCTGGAAAACGC | DrLGP2 core fragment |
5' RACE Outer Primer | CATGGCTACATGCTGACAGCCTA | DrLGP2 5' RACE |
5' RACE Inner Primer | CGCGGATCCACAGCCTACTGATGATCAGTCGATG | DrLGP2 5' RACE |
3' RACE Outer Primer | TACCGTCGTTCCACTAGTGATTT | DrLGP2 3' RACE |
3' RACE Inner Primer | CGCGGATCCTCCACTAGTGATTTCACTATAGG | DrLGP2 3' RACE |
LGP2-DEXDc-FW | ATCGAATTCATGGAGATCACTCTTAGATCAT | DrLGP2 antibody |
LGP2-DEXDc-BW | ATCGCGGCCGCGTGCTGGTCTACCAGGT | DrLGP2 antibody |
LGP2T-FW | ATGTGGCGTTACGTGGAGAA | DrLGP2T qRT-PCR |
LGP2T-BW | TCTGCAGCATCGGTGTGTAA | DrLGP2T qRT-PCR |
LGP2a-FW | GGCTGAACGGAAGAGGGAT | DrLGP2a qRT-PCR |
LGP2a-BW | TTCCTGATGTCTCCTCCGC | DrLGP2a qRT-PCR |
RIG-I-FW | TAAGGTGCTGAAGATGGCTC | RIG-I qRT-PCR |
RIG-I-BW | TGATGGTGTTCTGTCCGTTG | RIG-I qRT-PCR |
MDA5-FW | AGCCAGTTATCTGATCGGCG | MDA5 qRT-PCR |
MDA5-BW | TCAGCGATGTCCAAACCCTC | MDA5 qRT-PCR |
IFN-FW | TGCGTCTACTTGCGAATG | IFN qRT-PCR |
IFN-BW | GGCTTGGAAATGGTGTCT | IFN qRT-PCR |
Mx-FW | CTTGCGTGAGATGAGGTTGC | Mx qRT-PCR |
Mx-BW | TGTCTGGCGGCTCAGTAAGT | Mx qRT-PCR |
SHVV-N-FW | CCGCATCGGAAATCAAGCAG | N gene mRNA qRT-PCR |
SHVV-N-BW | GTTGACCGCTTGCCCAATTT | N gene mRNA qRT-PCR |
SHVV-G-FW | ACACCATACATGCCAGAGGC | G gene mRNA qRT-PCR |
SHVV-G-BW | GCCTCGCTGGGTATCCAAAT | G gene mRNA qRT-PCR |
SHVV-N-FW | CCGCATCGGAAATCAAGCAG | N gene cRNA qRT-PCR |
SHVV-N-BW | GTTGACCGCTTGCCCAATTT | N gene cRNA qRT-PCR |
SHVV-G-FW | ACACCATACATGCCAGAGGC | G gene cRNA qRT-PCR |
SHVV-G-BW | GCCTCGCTGGGTATCCAAAT | G gene cRNA qRT-PCR |
β-actin-FW | CACTGTGCCCATCTACGAG | β-actin qRT-PCR |
β-actin-BW | CCATCTCCTGCTCGAAGTC | β-actin qRT-PCR |
2.6. The Activation of the IFN-α mRNA Expression Was Associated with the Viral Replicative Intermediates
3. Discussion
4. Experimental Section
4.1. Cell, Virus and Infection
4.2. RNA Isolation and cDNA Synthesis
4.3. Amplification of Full-Length cDNAs of DrLGP2a and LGP2b by RACE
4.4. Sequence Analysis
4.5. Generation of Polyclonal DrLGP2-DEXDc Specific Antibody
4.6. Quantitative Real Time PCR (qRT-PCR) Assay for mRNA Expressions of Target Genes
4.7. Detection Protein Level of DrLGP2 by Western Blot
4.8. IFN-α Expression in ZF4 Cells Transfected with Total Cellular RNAs
Supplementary Materials
Acknowledgments
Author Contributions
Conflicts of Interest
References
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Wang, W.; Asim, M.; Yi, L.; Hegazy, A.M.; Hu, X.; Zhou, Y.; Ai, T.; Lin, L. Abortive Infection of Snakehead Fish Vesiculovirus in ZF4 Cells Was Associated with the RLRs Pathway Activation by Viral Replicative Intermediates. Int. J. Mol. Sci. 2015, 16, 6235-6250. https://doi.org/10.3390/ijms16036235
Wang W, Asim M, Yi L, Hegazy AM, Hu X, Zhou Y, Ai T, Lin L. Abortive Infection of Snakehead Fish Vesiculovirus in ZF4 Cells Was Associated with the RLRs Pathway Activation by Viral Replicative Intermediates. International Journal of Molecular Sciences. 2015; 16(3):6235-6250. https://doi.org/10.3390/ijms16036235
Chicago/Turabian StyleWang, Wenwen, Muhammad Asim, Lizhu Yi, Abeer M. Hegazy, Xianqin Hu, Yang Zhou, Taoshan Ai, and Li Lin. 2015. "Abortive Infection of Snakehead Fish Vesiculovirus in ZF4 Cells Was Associated with the RLRs Pathway Activation by Viral Replicative Intermediates" International Journal of Molecular Sciences 16, no. 3: 6235-6250. https://doi.org/10.3390/ijms16036235