Molecular Biology


    Protocols in Current Issue
    Characterize the Interaction of the DNA Helicase PriA with the Stalled DNA Replication Fork Using Atomic Force Microscopy
    Authors:  Yaqing Wang, Zhiqiang Sun, Piero R. Bianco and Yuri L. Lyubchenko, date: 03/05/2021, view: 936, Q&A: 0

    In bacteria, the restart of stalled DNA replication forks requires the DNA helicase PriA. PriA can recognize and remodel abandoned DNA replication forks, unwind DNA in the 3'-to-5' direction, and facilitate the loading of the helicase DnaB onto the DNA to restart replication. ssDNA-binding protein (SSB) is typically present at the abandoned forks,

    Chromatin Immunoprecipitation (ChIP) to Assess Histone Marks in Auxin-treated Arabidopsis thaliana Inflorescence Tissue
    Authors:  André Kuhn and Lars Østergaard, date: 12/05/2020, view: 1101, Q&A: 0
    [Abstract] Chromatin immunoprecipitation coupled with quantitative PCR (ChIP-qPCR) or high-throughput sequencing (ChIP-seq) has become the gold standard for the identification of binding sites of DNA binding proteins and the localization of histone modification on a locus-specific or genome-wide scale, respectively. ChIP experiments can be divided into seven ...
    In vitro Protein-DNA Binding Assay (AlphaScreen® Technology)
    Authors:  Mika Nomoto, Yasuomi Tada and Hironaka Tsukagoshi, date: 02/05/2019, view: 4573, Q&A: 0
    [Abstract] Identification of specific DNA binding sites of transcription factors is important in understanding their functions. Recent techniques allow us to investigate genome-wide in vivo binding positions by chromatin immunoprecipitation combined with high-throughput sequencing. However, to further explore the binding motifs of transcription ...
    Microtitre Plate Based Cell-SELEX Method
    Authors:  Munish Shorie and Harmanjit Kaur, date: 10/20/2018, view: 4178, Q&A: 0
    [Abstract] Aptamers have emerged as a novel category in the field of bioreceptors due to their wide applications ranging from biosensing to therapeutics. Several variations of their screening process, called SELEX have been reported which can yield sequences with desired properties needed for their final use. We report a facile microtiter plate-based ...
    ChIP-seq Experiment and Data Analysis in the Cyanobacterium Synechocystis sp. PCC 6803
    Authors:  Joaquín Giner-Lamia, Miguel A. Hernández-Prieto and Matthias E. Futschik, date: 06/20/2018, view: 8312, Q&A: 0
    [Abstract] Nitrogen is an essential nutrient for all living organisms. In cyanobacteria, a group of oxygenic photosynthetic bacteria, nitrogen homeostasis is maintained by an intricate regulatory network around the transcription factor NtcA. Although mechanisms controlling NtcA activity appear to be well understood, the sets of genes under its control (i.e. ...
    An Improved Method for Measuring Chromatin-binding Dynamics Using Time-dependent Formaldehyde Crosslinking
    [Abstract] Formaldehyde crosslinking is widely used in combination with chromatin immunoprecipitation (ChIP) to measure the locations along DNA and relative levels of transcription factor (TF)-DNA interactions in vivo. However, the measurements that are typically made do not provide unambiguous information about the dynamic properties of these ...
    Chromatin Affinity Purification (ChAP) from Arabidopsis thaliana Rosette Leaves Using in vivo Biotinylation System
    Authors:  Weronika Sura and Piotr A. Ziolkowski, date: 01/05/2018, view: 5694, Q&A: 0
    [Abstract] Chromatin Affinity Purification (ChAP) is widely used to study chromatin architecture and protein complexes interacting with DNA. Here we present an efficient method for ChAP from Arabidopsis thaliana rosette leaves, in which in vivo biotinylation system is used. The chromatin is digested by Micrococcal Nuclease (MNase), hence ...
    Chromatin Immunoprecipitation Experiments from Whole Drosophila Embryos or Larval Imaginal Discs
    [Abstract] Chromatin Immunoprecipitation coupled either to qPCR (qChIP) or high-throughput sequencing (ChIP-Seq) has been extensively used in the last decades to identify the DNA binding sites of transcription factors or the localization of various histone marks along the genome. The ChIP experiment generally includes 7 steps: collection of biological ...
    Micro-chromatin Immunoprecipitation (μChIP) Protocol for Real-time PCR Analysis of a Limited Amount of Cells
    Authors:  Sébastien Gillotin and François Guillemot, date: 06/20/2016, view: 6696, Q&A: 0
    [Abstract] Chromatin immunoprecipitation followed by deep sequencing (ChIP-Seq) is an important strategy to study gene regulation. When availability of cells is limited, however, it can be useful to focus on specific genes to investigate in depth the role of transcription factors or histone marks. Unfortunately, performing ChIP experiments to study ...
    MNase Digestion for Nucleosome Mapping in Neurospora
    Authors:  Cigdem Sancar, Gencer Sancar and Michael Brunner, date: 06/05/2016, view: 7448, Q&A: 0
    [Abstract] Digestion of chromatin by micrococcal nuclease MNase followed by high throughput sequencing allows us to determine the location and occupancy of nucleosomes on the genome. Here in this protocol we have described optimized conditions of MNase digestion of filamentous fungus Neurospora crassa chromatin without a requirement of a nuclear ...

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