Microbiology

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    Protocols in Current Issue
    Generation and in Planta Functional Analysis of Potato Virus Y mutants
    [Abstract] Potato virus Y (PVY), the type member of the genus Potyvirus (family Potyviridae), is the most widespread virus affecting potato and is included in the top five most economically detrimental plant viruses. Recently, the structure of the PVY virion has been determined by cryo-electron microscopy, which has opened the doors to ...
    Intestinal Enteroid Culture for Human Astroviruses
    Authors:  Irene A. Owusu, Carmen Mirabelli and Abimbola O. Kolawole, date: 07/20/2020, view: 238, Q&A: 0
    [Abstract] Human astroviruses (HAstV) are non-enveloped, positive-sense single stranded RNA viruses that typically cause gastroenteritis in children, the elderly and among immunocompromised individuals. Some HAstV species have also been implicated in neurological diseases. It is important to study these viruses to understand the pathogenesis and develop ...
    HIV-CRISPR: A CRISPR/Cas9 Screening Method to Identify Genes Affecting HIV Replication
    Authors:  Ferdinand Roesch and Molly OhAinle, date: 05/05/2020, view: 862, Q&A: 0
    [Abstract] Screening with CRISPR/Cas9 technology has already led to significant discoveries in the fields of cancer biology, cell biology and virology. Because of the relatively low false discovery rates and the ability to perform high-throughput, pooled approaches, it has rapidly become the assay of choice for screening studies, including whole-genome ...
    Purification of Rice Stripe Virus
    Authors:  Gang Lu, Min Yao, Yijun Zhou and Xiaorong Tao, date: 03/20/2020, view: 679, Q&A: 0
    [Abstract] Although many spherical and rod-shaped plant virus purification protocols are now available, only a few protocols on filamentous plant virus purification have been published. Here, we report a protocol for large-scale purification of Rice stripe virus (RSV) from RSV-infected rice tissues. RSV virions with high infectivity were first precipitated ...
    Expression and Purification of Adeno-associated Virus Virus-like Particles in a Baculovirus System and AAVR Ectodomain Constructs in E. coli
    [Abstract] Adeno-associated virus (AAV) is a promising gene therapy vector and the biophysical characterization of its interactions with host proteins is a critical foundation for engineering tissue targeting and immune escape. Presented here are protocols for the production of: (a) the outer protein shells (virus-like particles or VLPs) for serotype 2 ...
    Quantifying HIV-1-Mediated Gut CD4+ T Cell Death in the Lamina Propria Aggregate Culture (LPAC) Model
    Authors:  Stephanie M. Dillon, Kejun Guo, Moriah J. Castleman, Mario L. Santiago and Cara C. Wilson, date: 01/20/2020, view: 1072, Q&A: 0
    [Abstract] Gut CD4 T cells are major targets of HIV-1 and are massively depleted early during infection. To better understand the mechanisms governing HIV-1-mediated CD4 T cell death, we developed the physiologically-relevant Lamina Propria Aggregate Culture (LPAC) model. The LPAC model is ideal for studying CD4 T cell death induced by clinically-relevant ...
    Herpes Simplex Virus Type 1 Propagation, Titration and Single-step Growth Curves
    [Abstract] Given the endemic seroprevalence of herpes simplex viruses (HSV), its associated human diseases, and the emergence of acyclovir-resistant strains, there is a continuous need for better antiviral therapies. Towards this aim, identifying mechanistic details of how HSV-1 manipulates infected cells, how it modulates the immune responses, and how it ...
    Production of Quantum Dots-containing Influenza Virus Particles for Studying Viral Uncoating Processes
    Authors:  Chong Qin, Xiaowei Zhang and Zongqiang Cui, date: 09/20/2019, view: 1940, Q&A: 0
    [Abstract] The genome of influenza A virus (IAV) comprises eight pinlike genomic segments called vRNPs enclosed in viral capsid. During infection, uncoating is the key step for viral replication and represents an antiviral therapeutic target, but it is difficult to observe the transient and dynamic event in detail. Here, we report a protocol for production ...
    Image-based Quantification of Direct Cell-to-cell Transmission of Bovine Viral Diarrhea Virus
    Authors:  Fernando Merwaiss and Diego E. Alvarez, date: 08/05/2019, view: 1843, Q&A: 0
    [Abstract] Different viruses rely on direct cell-to-cell transmission to propagate infection within the infected host. Measuring this mode of transmission in cultured cells is often complicated by the contribution of cell free viruses to spread, and the difficulty to distinguish between primary infected cells that produce the virus and neighboring cells that ...
    Fluorescence Microscopy Assay to Measure HIV-1 Capsid Uncoating Kinetics in vitro
    [Abstract] The stability of the HIV-1 capsid and the spatiotemporal control of its disassembly, a process called uncoating, need to be finely tuned for infection to proceed. Biochemical methods for measuring capsid lattice disassembly in bulk are unable to resolve intermediates in the uncoating reaction. We have developed a single-particle fluorescence ...



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