AAV-Mediated HA-FAAH Gene Delivery

The N-terminally hemagglutinin (HA)-tagged mouse FAAH cDNA with 4 additional amino acids (Glu-Phe-Asp-Asn) between HA and the first codon of FAAH was cloned into the adeno-associated virus (AAV) plasmid containing the cytomegalovirus enhancer/chicken ß-actin (CAG) promoter [20 (link)], the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE), and the bovine growth hormone polyadenylation sequence (pA) flanked by AAV2 inverted terminal repeats. A transcriptional Stop cassette flanked by loxP sites was inserted downstream the CAG promoter to allow Cre recombinase-dependent transgene expression of HA-FAAH after injection into a Cre-expressing transgenic mouse line. Chimeric AAV serotype 1/2 vectors were produced and genomic titers determined using the Applied Biosystems ABI 7500 real time PCR cycler as described before [22 (link)].

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Zimmermann T., Bartsch J.C., Beer A., Lomazzo E., Guggenhuber S., Lange M.D., Bindila L., Pape H.C, & Lutz B. (2018). Impaired anandamide/palmitoylethanolamide signaling in hippocampal glutamatergic neurons alters synaptic plasticity, learning, and emotional responses. Neuropsychopharmacology, 44(8), 1377-1388.

Publication 2018

ABI 7500 real time PCR cycler

Actin Adeno associated virus Adeno associated virus 1 Amino acids Asp glu Asp phe Bovine growth hormone Cdna Chicken Chimeric Codon Cre recombinase Cytomegalovirus Genomic Glu asp Hemagglutinin Inverted terminal repeats Mouse Phe glu Plasmid Polyadenylation Real time pcr Regulatory element Transcriptional Transgene Transgenic mouse Vectors Woodchuck hepatitis virus

Corresponding Organization : Johannes Gutenberg University Mainz

Other organizations : University of Münster

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Variable analysis

independent variables

The N-terminally hemagglutinin (HA)-tagged mouse FAAH cDNA with 4 additional amino acids (Glu-Phe-Asp-Asn) between HA and the first codon of FAAH

dependent variables

Not explicitly mentioned

control variables

The adeno-associated virus (AAV) plasmid containing the cytomegalovirus enhancer/chicken ß-actin (CAG) promoter, the woodchuck hepatitis virus posttranscriptional regulatory element (WPRE), and the bovine growth hormone polyadenylation sequence (pA) flanked by AAV2 inverted terminal repeats
The transcriptional Stop cassette flanked by loxP sites inserted downstream the CAG promoter to allow Cre recombinase-dependent transgene expression of HA-FAAH
Chimeric AAV serotype 1/2 vectors

controls

Positive control: Not explicitly mentioned
Negative control: Not explicitly mentioned

Annotations

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