Molecular Biology


    Protocols in Current Issue
    Quantification of Extracellular Double-stranded RNA Uptake and Subcellular Localization Using Flow Cytometry and Confocal Microscopy
    Authors:  Tan A Nguyen, Lachlan Whitehead and Ken C Pang, date: 06/20/2018, view: 68, Q&A: 0
    [Abstract] Double-stranded RNA is a potent pathogen-associated molecular pattern (PAMP) produced as a by-product of viral replication and a well-known hallmark of viral infection. Viral dsRNAs can be released from infected cells into the extracellular space and internalized by neighboring cells via endocytosis. Mammals possess multiple pattern recognition ...
    Characterizing the Transcriptional Effects of Endolysin Treatment on Established Biofilms of Staphylococcus aureus
    [Abstract] Biofilms are the most common lifestyle of bacteria in both natural and human environments. The organized structure of these multicellular communities generally protects bacterial cells from external challenges, thereby enhancing their ability to survive treatment with antibiotics or disinfectants. For this reason, the search for new antibiofilm ...
    Dual-probe RNA FRET-FISH in Yeast
    Authors:  Gable M. Wadsworth, Rasesh Y. Parikh and Harold D. Kim, date: 06/05/2018, view: 298, Q&A: 0
    [Abstract] mRNA Fluorescence In Situ Hybridization (FISH) is a technique commonly used to profile the distribution of transcripts in cells. When combined with the common single molecule technique Fluorescence Resonance Energy Transfer (FRET), FISH can also be used to profile the co-expression of nearby sequences in the transcript to measure ...
    Single-probe RNA FISH in Yeast
    Authors:  Gable M. Wadsworth, Rasesh Y. Parikh and Harold D. Kim, date: 06/05/2018, view: 200, Q&A: 0
    [Abstract] Quantitative profiling of mRNA expression is an important part of understanding the state of a cell. The technique of RNA Fluorescence In Situ Hybridization (FISH) involves targeting an RNA transcript with a set of 40 complementary fluorescently labeled DNA oligonucleotide probes. However, there are many circumstances such as transcripts ...
    Ectopic Gene Expression in Macrophages Using in vitro Transcribed mRNA
    Authors:  Pallavi Chandra and Jennifer A. Philips, date: 05/20/2018, view: 361, Q&A: 0
    [Abstract] Macrophages are immune cells that contribute to host defense through various mechanisms including phagocytosis and antigen presentation. Their antimicrobial capacity is subverted by clinically important intracellular pathogens such as Mycobacterium tuberculosis. The study of host-pathogen interactions using these cells is therefore of ...
    A Modified Low-quantity RNA-Seq Method for Microbial Community and Diversity Analysis Using Small Subunit Ribosomal RNA
    Authors:  Yong-Wei Yan, Ting Zhu, Bin Zou and Zhe-Xue Quan, date: 05/05/2018, view: 246, Q&A: 0
    [Abstract] We propose a modified RNA-Seq method for small subunit ribosomal RNA (SSU rRNA)-based microbial community analysis that depends on the direct ligation of a 5’ adaptor to RNA before reverse-transcription. The method requires only a low-input quantity of RNA (10-100 ng) and does not require a DNA removal step. Using this method, we could obtain more ...
    Purification of Total RNA from DSS-treated Murine Tissue via Lithium Chloride Precipitation
    Authors:  Emilie Viennois, Anika Tahsin and Didier Merlin, date: 05/05/2018, view: 401, Q&A: 0
    [Abstract] We have developed a protocol to purify RNA from DSS (Dextran Sulfate Sodium)-treated mouse tissues. This method, which prevents downstream inhibition of q-RT-PCR observed in DSS-treated tissues, relies on successive precipitations with lithium chloride.
    Assessing the Efficacy of Small Molecule Inhibitors in a Mouse Model of Persistent Norovirus Infection
    Authors:  Jana Van Dycke, Johan Neyts and Joana Rocha-Pereira, date: 05/05/2018, view: 389, Q&A: 0
    [Abstract] Human norovirus is the most common cause of acute gastroenteritis worldwide, resulting in estimated mortality of ~210,000 each year, of whom most are children under the age of five. However, norovirus can infect people of all age groups. There is a risk of prolonged infection in children, the elderly and patients who are immunocompromised. To ...
    Detecting the Interaction of Double-stranded RNA Binding Protein, Viral Protein and Primary miRNA Transcript by Co-immunoprecipitation in planta
    Authors:  Lijia Zheng, Chao Zhang, Jianguo Wu and Yi Li, date: 05/05/2018, view: 277, Q&A: 0
    [Abstract] MicroRNAs (miRNAs) play important roles in plant growth, development, and response to infection by microbes. Double-stranded RNA binding protein 1 (DRB1) facilitates the processing of primary miRNA transcripts into mature miRNAs. Recently, we found that NS3 protein encoded by rice stripe virus (RSV) associates with DRB1 and promotes miRNA ...
    Generation of microRNA Sponge Library
    Author:  Sebastian Herzog, date: 04/20/2018, view: 328, Q&A: 0
    [Abstract] This protocol describes the generation and functional validation of microRNA (miRNA) sponge or decoy constructs. When expressed from a strong promoter, these transcripts can sequester specific miRNA:RISC complexes, thereby resulting in a derepression of endogenous target mRNA. Hence, cells expressing such sponges display a partial or full miRNA ...