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ARS Home » Plains Area » Kerrville, Texas » Knipling-Bushland U.S. Livestock Insects Research Laboratory » LAPRU » Research » Publications at this Location » Publication #321453
Title: Identification, characterization and use of two tick promoters for construction of a dual luciferase reporter vector

Author
item Tuckow, Alexander
item Temeyer, Kevin

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 2/6/2015
Publication Date: N/A
Citation: N/A

Interpretive Summary:

Technical Abstract: Dual luciferase reporter systems are valuable tools for functional genomic studies, but have not previously been developed for use in tick cell culture. We evaluated expression of available luciferase constructs in tick cell cultures derived from Rhipicephalus (Boophilus) microplus, an important vector of bovine babesiosis and anaplasmosis. Commercial promoters were evaluated for transcriptional activity driving luciferase expression in the tick cell lines. The human PGK (phosphoglycerate kinase) promoter resulted in detectable firefly luciferase activity within 2 days post-transfection of the R. microplus cell line BmE26, with maximal activity at 5 days post-transfection. Several other promoters were weaker or inactive in the tick cells, prompting identification and assessment of transcriptional activity of the homologous ribosomal protein L4 (rpL4, GenBank accession: KM516205) and elongation factor 1a (EF-1a, GenBank accession: KM516204) promoters cloned from R. microplus. Evaluation of luciferase expression driven by various promoters in tick cell culture resulted in selection of the R. microplus RPL4 promoter and the human PGK promoter driving transcription of sequences encoding modified firefly and NanoLuc® luciferases for construction of a dual luciferase reporter system for use in tick cell culture.