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J. Craig Venter Institute

4 ARTICLES PUBLISHED IN JoVE

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Immunology and Infection

Isolation and Genome Analysis of Single Virions using 'Single Virus Genomics'
Lisa Zeigler Allen 1, Thomas Ishoey 1, Mark A. Novotny 1, Jeffrey S. McLean 1, Roger S. Lasken 1, Shannon J. Williamson 1
1Department of Microbial and Environmental Genomics, The J. Craig Venter Institute

Single Virus Genomics (SVG) is a method to isolate and amplify the genomes of single virons. Viral suspensions of a mixed assemblage are sorted using flow cytometry onto a microscope slide with discrete wells containing agarose, thereby capturing the virion and reducing genome shearing during downstream processing. Whole genome amplification is achieved using multiple displacement amplification (MDA) resulting in genomic material that is suitable for sequencing.

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Biology

The Green Monster Process for the Generation of Yeast Strains Carrying Multiple Gene Deletions
Yo Suzuki 1, Jason Stam 1, Mark Novotny 2, Nozomu Yachie 3, Roger S. Lasken 2, Frederick P. Roth 3,4
1Department of Synthetic Biology and Bioenergy, J. Craig Venter Institute, 2Department of Microbial and Environmental Genomics, J. Craig Venter Institute, 3Donnelly Centre & Department of Molecular Genetics, University of Toronto, 4Lunenfeld Research Institute, Mt Sinai Hospital

The Green Monster method enables the rapid assembly of multiple deletions marked with a reporter gene encoding green fluorescent protein. This method is based on driving yeast strains through repeated cycles of sexual assortment of deletions and fluorescence-based enrichment of cells carrying more deletions.

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Immunology and Infection

Generation of Escape Variants of Neutralizing Influenza Virus Monoclonal Antibodies
Paul E. Leon 1,2, Teddy John Wohlbold 1,2, Wenqian He 1,2, Mark J. Bailey 1,2, Carole J. Henry 3, Patrick C. Wilson 3, Florian Krammer 1, Gene S. Tan 1
1Department of Microbiology, Icahn School of Medicine at Mount Sinai, 2Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, 3The Department of Medicine, Section of Rheumatology, The Knapp Center for Lupus and Immunology Research, The University of Chicago

We describe a method by which we identify critical residues required for the binding of human or murine monoclonal antibodies that target the viral hemagglutinin of influenza A viruses. The protocol can be adapted to other virus surface glycoproteins and their corresponding neutralizing antibodies.

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Immunology and Infection

A Method to Assess Fc-mediated Effector Functions Induced by Influenza Hemagglutinin Specific Antibodies
Mark J. Bailey 1,2, Felix Broecker 1, Paul E. Leon 1,2, Gene S. Tan 3
1Department of Microbiology, Icahn School of Medicine at Mount Sinai, 2Graduate School of Biomedical Sciences, Icahn School of Medicine at Mount Sinai, 3Department of Infectious Disease, J. Craig Venter Institute

We describe a method to measure the activation of Fc-mediated effector functions by antibodies that target the influenza virus hemagglutinin. This assay can also be adapted to assess the ability of monoclonal antibodies or polyclonal sera targeting other viral surface glycoproteins to induce Fc-mediated immunity.

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