Microbiology

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    Protocols in Current Issue
    Quantification of Hepatitis B Virus Covalently Closed Circular DNA in Infected Cell Culture Models by Quantitative PCR
    Authors:  Bingqian Qu and Stephan Urban, date: 04/05/2019, view: 500, Q&A: 0
    [Abstract] Persistence of the human hepatitis B virus (HBV) requires the maintenance of covalently closed circular (ccc)DNA, the episomal genome reservoir in nuclei of infected hepatocytes. cccDNA elimination is a major aim in future curative therapies currently under development. In cell culture based in vitro studies, both hybridization- and ...
    Quantification of Queuosine Modification Levels in tRNA from Human Cells Using APB Gel and Northern Blot
    Authors:  Zaneta Matuszek and Tao Pan, date: 03/20/2019, view: 510, Q&A: 0
    [Abstract] Queuosine (Q) is a hypermodified base in the wobble anticodon position of tRNAs coding for the amino acids Tyr, His, Asn, and Asp. tRNA Q-modification is introduced by a queuine tRNA-ribosyltransferase (TGT) that replaces the guanine base at G34 at these tRNAs with the modified base. tRNA Q-modification is widely distributed among prokaryotic and ...
    Isolation of Thylakoid Membranes from the Cyanobacterium Synechocystis sp. PCC 6803 and Analysis of Their Photosynthetic Pigment-protein Complexes by Clear Native-PAGE
    Authors:  Josef Komenda, Vendula Krynická and Tomas Zakar, date: 01/05/2019, view: 1044, Q&A: 0
    [Abstract] Cyanobacteria represent a frequently used model organism for the study of oxygenic photosynthesis. They belong to prokaryotic microorganisms but their photosynthetic apparatus is quite similar to that found in algal and plant chloroplasts. The key players in light reactions of photosynthesis are Photosystem I and Photosystem II complexes (PSI and ...
    Detection of D-glutamate Production from the Dual Function Enzyme, 4-amino-4-deoxychorismate Lyase/D-amino Acid Transaminase, in Mycobacterium smegmatis
    Authors:  Helen K. Opel-Reading, Roman Mortuza and Kurt L. Krause, date: 01/05/2019, view: 790, Q&A: 0
    [Abstract] D-amino acid transaminase (D-AAT) is able to synthesize both D-glutamate and D-alanine, according to the following reaction: D-alanine + α-ketoglutarate ⇌ D-glutamate + pyruvate. These two D-amino acids are essential components of the peptidoglycan layer of bacteria. In our recently published work, MSMEG_5795 from Mycobacterium smegmatis ...
    Analysis of DNA Exchange Using Thymidine Analogs (ADExTA) in Trypanosoma cruzi
    [Abstract] Trypanosoma cruzi is a protozoan parasite belonging to the Trypanosomatidae family. Although the trypanosomatids multiply predominantly by clonal generation, the presence of DNA exchange in some of them has been puzzling researchers over the years, mainly because it may represent a novel form that these organisms use to gain variability. ...
    Extraction and Analysis of Bacterial Teichoic Acids
    Authors:  Kelvin Kho and Timothy C. Meredith, date: 11/05/2018, view: 1182, Q&A: 0
    [Abstract] Teichoic acids (TA) are anionic polymers comprised of polyol phosphate repeat units that are abundant in the cell wall of Gram-positive bacteria. Both wall teichoic acid (WTA) and lipoteichoic acid (LTA) play important roles in regulating cell wall remodeling as well as conferring antibiotic resistance. To analyze TA, we describe a polyacrylamide ...
    Kinetic Characterization of the Shigella Type Three Secretion System ATPase Spa47 Using α-32P ATP
    Authors:  Heather B. Case and Nicholas E. Dickenson, date: 11/05/2018, view: 993, Q&A: 0
    [Abstract] ATPases represent a diverse class of enzymes that utilize ATP hydrolysis to support critical biological functions such as driving ion pumps, providing mechanical work, unfolding/folding proteins, and supporting otherwise thermodynamically unfavorable chemical reactions. We have recently shown that the Shigella protein Spa47 is an ATPase ...
    Identifying Protein Interactions with Histone Peptides Using Bio-layer Interferometry
    Authors:  Bingbing Ren, Ahmed Mahmoud Mohammed Sayed, Hwei Ling Tan, Yu Keung Mok and Ee Sin Chen, date: 09/20/2018, view: 1596, Q&A: 0
    [Abstract] Histone post-translational modifications (PTMs) regulate numerous cellular processes, including gene transcription, cell division, and DNA damage repair. Most histone PTMs affect the recruitment or exclusion of reader proteins from chromatin. Here, we present a protocol to measure affinity and interaction kinetics between histone peptides and the ...
    Extraction and Quantification of Polyphosphate (polyP) from Gram-negative Bacteria
    Authors:  Jan-Ulrik Dahl, Lihan Xie and Ursula Jakob, date: 09/20/2018, view: 1209, Q&A: 0
    [Abstract] Polyphosphate (polyP), a universally conserved biomolecule, is composed of up to 1,000 phosphate monomers linked via phosphoanhydride bonds. Reaching levels in bacteria that are in the high nmoles per mg protein range, polyP plays important roles in biofilm formation and colonization, general stress protection and virulence. Various protocols for ...
    Selective Isolation of Retroviruses from Extracellular Vesicles by Intact Virion Immunoprecipitation
    Authors:  Tyler Milston Renner, Kasandra Bélanger and Marc-André Langlois, date: 09/05/2018, view: 1234, Q&A: 0
    [Abstract] There exists a wide variety of techniques to isolate and purify viral particles from cell culture supernatants. However, these techniques vary greatly in ease of use, purity, yield and impact on viral structural integrity. Most importantly, it is becoming evident that secreted extracellular vesicles (EVs) co-purify with retroviruses using nearly ...



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