Robert C. Byrd Biotechnology Science Center

3 ARTICLES PUBLISHED IN JoVE

Immunology and Infection

Identification of Novel Genes Associated with Alginate Production in Pseudomonas aeruginosa Using Mini-himar1 Mariner Transposon-mediated Mutagenesis

T. Ryan Withers¹, Yeshi Yin¹, Hongwei D. Yu¹

¹Department of Biochemistry and Microbiology, Joan C. Edwards School of Medicine, Marshall University

Here we describe a protocol using the mini-himar1 mariner transposon-mediated mutagenesis for generating a high-density insertion mutant library to screen, isolate and identify novel alginate regulators in the prototypic Pseudomonas aeruginosa strain PAO1.

Immunology and Infection

Generation of In-Frame Gene Deletion Mutants in Pseudomonas aeruginosa and Testing for Virulence Attenuation in a Simple Mouse Model of Infection

Meagan E. Valentine¹, Brandon D. Kirby¹, Hongwei D. Yu^1,2

¹Progenesis Technologies, LLC, ²Department of Biomedical Sciences, Pediatrics, Joan C. Edwards School of Medicine at Marshall University

Here, we describe a simple and reproducible protocol of mouse model of infection to evaluate the attenuation of the genetically modified strains of Pseudomonas aeruginosa in comparison to the United States Food and Drug Administration (FDA)-approved Escherichia coli for commercial applications.

Immunology and Infection

Culture of Small Colony Variant of Pseudomonas aeruginosa and Quantitation of its Alginate

Roy Al Ahmar^*1, Brandon D. Kirby^*2, Hongwei D. Yu^1,2,3

¹Department of Biomedical Sciences, Joan C. Edwards School of Medicine, Marshall University, ²Progenesis Technologies LLC, Robert C. Byrd Biotechnology Science Center, ³Department of Pediatrics, Joan C. Edwards School of Medicine, Marshall University

Here, we describe a growth condition to culture the small colony variant of Pseudomonas aeruginosa. We also describe two separate methods for the detection and quantitation of the exopolysaccharide alginate produced by P. aeruginosa using a traditional uronic acid carbazole assay and an alginate-specific monoclonal antibody (mAb) based ELISA.