Plasmid construction for ZIKV and related viral proteins

The cDNA coding for each ZIKV protein, including prM, NS4A and NS4A-Flag (with a C-terminal Flag epitope tag), was PCR amplified from ZG-01 strain viral cDNA as a template and cloned into pFN11A (Promega, San Luis Obispo, CA) or pcDNA3.1 vectors. Plasmids including pcDNA3.1-DENV NS4A, -IVA PB1-F2, -MAVS, -RIG-I (N), pFN10A (ACT)-RIG-I, -MAVS, -TBK1, and-IKKε were constructed as previously described [22 (link)]. Truncated forms of MAVS (aa 1–77, 74–173 and 174–540) with N-terminal Flag epitope tags were amplified from the full-length template and cloned into the pcDNA3.1 vector. The pIFN-β-luc reporter plasmid was constructed by cloning a 125 bp fragment of the IFN-β promoter into the pGL3-Basic vector using the NheI and HindIII sites upstream of the luciferase reporter gene [8 (link), 22 (link)]. All constructs were verified by DNA sequencing. All primers used for plasmid construction are listed in Additional file 3: Table S3.

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Hu Y., Dong X., He Z., Wu Y., Zhang S., Lin J., Yang Y., Chen J., An S., Yin Y., Shen Z., Zeng G., Tian H., Cai J., Yang Y., Guan H., Wu J., Li M, & Zhu X. (2019). Zika virus antagonizes interferon response in patients and disrupts RIG-I–MAVS interaction through its CARD-TM domains. Cell & Bioscience, 9, 46.

Publication 2019

pFN11A

Cdna Epitope Luciferase Mavs Pgl3 Plasmid Primers Promega Protein Reporter gene Rig i Strain Tbk1 Vector Zikv

Corresponding Organization :

Other organizations : Sun Yat-sen University, Guangzhou Medical University, The First Affiliated Hospital, Sun Yat-sen University

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Variable analysis

independent variables

CDNA coding for each ZIKV protein, including prM, NS4A and NS4A-Flag (with a C-terminal Flag epitope tag)
Truncated forms of MAVS (aa 1–77, 74–173 and 174–540) with N-terminal Flag epitope tags

dependent variables

IFN-β promoter activity (as measured by the pIFN-β-luc reporter plasmid)

control variables

PcDNA3.1-DENV NS4A, -IVA PB1-F2, -MAVS, -RIG-I (N), pFN10A (ACT)-RIG-I, -MAVS, -TBK1, and-IKKε plasmids

positive controls

Full-length MAVS template

negative controls

Not explicitly mentioned

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