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URL: http://www.addgene.org/110916

Proper Citation: RRID:Addgene_110916

Insert Name: unc-129 promoter

Organism: Caenorhabditis elegans

Bacterial Resistance: Ampicillin

Defining Citation: PMID:30401765

Vector Backbone Description: Backbone Size:2617; Vector Backbone:pUC; Vector Types:Bacterial Expression; Bacterial Resistance:Ampicillin

Comments: We used Herculase II DNA polymerase and primers engineered with restriction sites to amplify and clone the 1.4 Kb strn-1 (C16E9.2) cDNA (minus its stop codon) from KG#720 rab-3::strn-1-FLAG into Nhe I/ Kpn I cut KG#367 (unc-129::___-GFP expression vector; 7.2 Kb) such that it is in-frame with the GFP. Transformed into XL1-Blue electrocompetent cells. Did Qiagen preps on 2 clones to find one with correct size insert for sequence verification. Verified sequence. Features of the construct: The unc-129:: promoter drives expression in a subset of 9 DA/ DB cholinergic motor neurons. AAAA precedes the start codon to provide a consensus ribosome binding site. A non-structured linker (SSGSSG) replaces the strn-1 stop codon and is immediately followed by the Fire vector GFP with 3 artificial introns (there are also introns in the 5' and 3' UTR in this vector). 2 bases are added to maintain the reading frame with the downstream GFP when cloning into the Kpn I site (this encodes an Ala).

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