Plasmid Construction and Antibody Generation

The full-length porcine NME1 cDNA was amplified and inserted into pcDNA^TM3.1/myc-His(-)A vector (Invitrogen) to generate a plasmid expressing Myc-tagged NME1 (Myc-NME1). A series of plasmids expressing Flag-tagged viral proteins were constructed as described previously⁵². HA-tagged p53 plasmid (HA-p53) was constructed by inserting full-length p53 cDNA into pCAGGs vector (including a C-terminal HA tag). Flag-p53, p53-Luc reporter plasmids and control plasmid Renilla luciferase pRL-TK were kindly provided by Zhiyong Ma (Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences, China)^{53 (link)}. All the expressing plasmids used in this study were analyzed and verified by DNA sequencing.
Commercial antibodies included: mouse monoclonal anti-Myc, monoclonal anti-Flag, rabbit polyclonal anti-NME1, and mouse monoclonal anti-β-actin (Santa Cruz Biotechnology); mouse monoclonal anti-hemagglutinin (HA) (BioLegend); mouse monoclonal anti-Flag (Sigma); rabbit polyclonal anti-eukaryotic translation initiation factor 4 gamma (eIF4G) (Abcam); rabbit polyclonal anti-LC3B (Sigma); rabbit monoclonal anti-ATG12 (Cell signaling technology); rabbit polyclonal anti-VP1 antibody was prepared by our laboratory as described previously^{48 (link)}.

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Feng H.H., Zhu Z.X., Cao W.J., Yang F., Zhang X.L., Du X.L., Zhang K.S., Liu X.T, & Zheng H.X. (2018). Foot-and-mouth disease virus induces lysosomal degradation of NME1 to impair p53-regulated interferon-inducible antiviral genes expression. Cell Death & Disease, 9(9), 885.

Publication 2018

pcDNATM3.1/myc-His(-)A vector mouse monoclonal anti-Myc monoclonal anti-Flag mouse monoclonal anti-β-actin mouse monoclonal anti-hemagglutinin (HA) mouse monoclonal anti-Flag rabbit polyclonal anti-LC3B

Actin Anti antibody Antibodies Cdna Chinese Eukaryotic translation initiation factor 4 gamma Hemagglutinin Mouse Nme1 Plasmids Porcine Rabbit Renilla luciferase Vector Viral proteins

Corresponding Organization :

Other organizations : Chinese Academy of Agricultural Sciences, Lanzhou Veterinary Research Institute

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Variable analysis

independent variables

Myc-tagged NME1 (Myc-NME1)
Flag-tagged viral proteins
HA-tagged p53 (HA-p53)
Flag-p53

dependent variables

Not explicitly mentioned

control variables

PcDNA™3.1/myc-His(-)A vector (Invitrogen)
PCAGGs vector
P53-Luc reporter plasmids
Renilla luciferase pRL-TK

controls

Positive control: Renilla luciferase pRL-TK
Negative control: Not explicitly mentioned

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