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ARS Home » Plains Area » Kerrville, Texas » Knipling-Bushland U.S. Livestock Insects Research Laboratory » LAPRU » Research » Research Project #429222

Research Project: Identification of Novel Vaccine Candidates against Transmission of Arthropod-Borne Pathogens

Location: Livestock Arthropod Pests Research

Project Number: 3094-32000-036-51-S
Project Type: Non-Assistance Cooperative Agreement

Start Date: Aug 1, 2015
End Date: Jul 31, 2020

Determine the role of inward rectifying potassium (Kir) channels in the arthropod salivary gland. Determine salivary gland and gut genes that are differentially expressed during arthropod feeding that may serve as alternate target sites for vaccine development.

Objective 1: Test if knockdown of Kir channel constructs in Drosophila salivary glands reduces the feeding efficacy of the flies; explore the role of Kir channels in the salivary gland using the GAL4/UAS system; cross fly strains expressing the GAL4/UAS promoter or the dsRNA against Kir channels and screen the F1 progeny for the knockdown phenotype; utilize a Drosophila feeding assay (objective 1A) in the knockdown and control lines to determine if Kir channels constitute a critical ion channel for proper feeding; move the study into horn fly and tick salivary glands by employing the Ramsay bioassay with isolated salivary glands that characterizes the ion and fluid secretion rates across the membrane, which are biomarkers of salivation abilities. Objective 2: Collect multiple time points of feeding ticks and extract the salivary gland from each adult tick to determine the feeding-associated differential gene expression in ticks; utilize established methods for RNA isolation, transcriptome sequencing and analysis; identify the top 4 genes that are differentially expressed and begin the iterative process of genetically manipulating the gene expression in Drosophila, determine the influence on feeding using the feeding assay, and perform the Ramsey assay using isolated salivary glands to determine the influence of the gene to the animal. Express two transcripts proved crucial to feeding, salivary gland function, or osmoregulation as recombinant proteins in yeast to begin vaccine studies.