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Methods in Molecular Biology (Clifton,... 1997
Review
Topics: Base Sequence; DNA; DNA Footprinting; Molecular Sequence Data
PubMed: 9407525
DOI: 10.1385/0-89603-447-X:23 -
Nature Methods Mar 2016The advent of DNA footprinting with DNase I more than 35 years ago enabled the systematic analysis of protein-DNA interactions, and the technique has been instrumental... (Review)
Review
The advent of DNA footprinting with DNase I more than 35 years ago enabled the systematic analysis of protein-DNA interactions, and the technique has been instrumental in the decoding of cis-regulatory elements and the identification and characterization of transcription factors and other DNA-binding proteins. The ability to analyze millions of individual genomic cleavage events via massively parallel sequencing has enabled in vivo DNase I footprinting on a genomic scale, offering the potential for global analysis of transcription factor occupancy in a single experiment. Genomic footprinting has opened unique vistas on the organization, function and evolution of regulatory DNA; however, the technology is still nascent. Here we discuss both prospects and challenges of genomic footprinting, as well as considerations for its application to complex genomes.
Topics: Algorithms; Chromosome Mapping; DNA; DNA Footprinting; Genome, Human; High-Throughput Nucleotide Sequencing; Humans
PubMed: 26914205
DOI: 10.1038/nmeth.3768 -
Nature Protocols Dec 2021Precise control of gene expression requires the coordinated action of multiple factors at cis-regulatory elements. We recently developed single-molecule footprinting to... (Review)
Review
Precise control of gene expression requires the coordinated action of multiple factors at cis-regulatory elements. We recently developed single-molecule footprinting to simultaneously resolve the occupancy of multiple proteins including transcription factors, RNA polymerase II and nucleosomes on single DNA molecules genome-wide. The technique combines the use of cytosine methyltransferases to footprint the genome with bisulfite sequencing to resolve transcription factor binding patterns at cis-regulatory elements. DNA footprinting is performed by incubating permeabilized nuclei with recombinant methyltransferases. Upon DNA extraction, whole-genome or targeted bisulfite libraries are prepared and loaded on Illumina sequencers. The protocol can be completed in 4-5 d in any laboratory with access to high-throughput sequencing. Analysis can be performed in 2 d using a dedicated R package and requires access to a high-performance computing system. Our method can be used to analyze how transcription factors cooperate and antagonize to regulate transcription.
Topics: Animals; Cell Nucleus; DNA; DNA Footprinting; DNA Modification Methylases; Gene Expression Regulation; Gene Library; Genome; High-Throughput Nucleotide Sequencing; Humans; Mice; Mouse Embryonic Stem Cells; Nucleosomes; RNA Polymerase II; Sequence Analysis, DNA; Single Molecule Imaging; Software; Transcription Factors
PubMed: 34773120
DOI: 10.1038/s41596-021-00630-1 -
Methods in Molecular Biology (Clifton,... 1999
Review
Topics: Animals; Cell Extracts; DNA Footprinting; DNA-Binding Proteins; Xenopus laevis
PubMed: 10503235
DOI: 10.1385/1-59259-678-9:187 -
Methods in Molecular Biology (Clifton,... 2003
Topics: Cloning, Molecular; DNA Footprinting; DNA Methylation; Electrophoresis, Polyacrylamide Gel; Genetic Techniques; Plasmids; Polymerase Chain Reaction; Precipitin Tests
PubMed: 12777724
DOI: 10.1385/1-59259-329-1:117 -
Molekuliarnaia Biologiia 2018Ligand binding influences the dynamics of the DNA helix in both the binding site and adjacent regions. This, in particular, is reflected in the changing pattern of...
Ligand binding influences the dynamics of the DNA helix in both the binding site and adjacent regions. This, in particular, is reflected in the changing pattern of cleavage of complexes under the action of ultrasound. The specificity of ultrasound-induced cleavage of the DNA sugar-phosphate backbone was studied in actinomycin D (AMD) complexes with double-stranded DNA restriction fragments. After antibiotic binding, the cleavage intensity of phosphodiester bonds between bases was shown to decrease at the chromophore intercalation site and to increase in adjacent positions. The character of cleavage depended on the sequences flanking the binding site and the presence of other AMD molecules bound in the close vicinity. A comparison of ultrasonic and DNase I cleavage patterns of AMD-DNA complexes provided more detail on the local conformation and dynamics of the DNA double helix in both binding site and adjacent regions. The results pave the way for developing a novel approach to studies of the nucleotide sequence dependence of DNA conformational dynamics and new techniques to identify functional genome regions.
Topics: Base Sequence; Binding Sites; DNA; DNA Footprinting; DNA-Binding Proteins; Dactinomycin; Deoxyribonuclease I; Gene Expression; Intercalating Agents; Ligands; Nucleic Acid Conformation; Ultrasonic Waves
PubMed: 30113037
DOI: 10.1134/S0026898418040067 -
Methods (San Diego, Calif.) Jun 2007Footprinting is a simple method for assessing the sequence selectivity of DNA-binding ligands. The method is based on the ability of the ligand to protect DNA from... (Review)
Review
Footprinting is a simple method for assessing the sequence selectivity of DNA-binding ligands. The method is based on the ability of the ligand to protect DNA from cleavage at its binding site. This review describes the use of DNase I and hydroxyl radicals, the most commonly used footprinting probes, in footprinting experiments. The success of a footprinting experiment depends on using an appropriate DNA substrate and we describe how these can best be chosen or designed. Although footprinting was originally developed for assessing a ligand's sequence selectivity, it can also be employed to estimate the binding strength (quantitative footprinting) and to assess the association and dissociation rate constants for slow binding reactions.
Topics: Base Sequence; Binding Sites; DNA; DNA Footprinting; Deoxyribonuclease I; Hydroxyl Radical; Kinetics; Ligands; Sensitivity and Specificity; Substrate Specificity
PubMed: 17472895
DOI: 10.1016/j.ymeth.2007.01.002 -
Nature Methods Oct 2015Regulatory regions harbor multiple transcription factor (TF) recognition sites; however, the contribution of individual sites to regulatory function remains challenging...
Regulatory regions harbor multiple transcription factor (TF) recognition sites; however, the contribution of individual sites to regulatory function remains challenging to define. We describe an approach that exploits the error-prone nature of genome editing-induced double-strand break repair to map functional elements within regulatory DNA at nucleotide resolution. We demonstrate the approach on a human erythroid enhancer, revealing single TF recognition sites that gate the majority of downstream regulatory function.
Topics: Base Sequence; Binding Sites; Carrier Proteins; DNA Breaks, Double-Stranded; DNA Footprinting; DNA Repair; Enhancer Elements, Genetic; Erythrocytes; Erythropoiesis; Genome, Human; Genomics; Humans; Mutation; Nuclear Proteins; Regulatory Sequences, Nucleic Acid; Repressor Proteins; Transcription Factors
PubMed: 26322838
DOI: 10.1038/nmeth.3554 -
Methods in Molecular Biology (Clifton,... 1997
Review
Topics: Antineoplastic Agents; Base Sequence; DNA; DNA Footprinting; Molecular Sequence Data; Transcription, Genetic
PubMed: 9407532
DOI: 10.1385/0-89603-447-X:127 -
Methods in Molecular Biology (Clifton,... 2010Footprinting is a method for determining the sequence selectivity of DNA-binding compounds in which ligands protect DNA from cleavage at their binding sites....
Footprinting is a method for determining the sequence selectivity of DNA-binding compounds in which ligands protect DNA from cleavage at their binding sites. Footprinting templates are typically 50-200 base pairs long, and DNase I is the most commonly used nuclease for these experiments. This chapter describes the preparation and labelling of suitable DNA footprinting substrates, the footprinting experiment itself, and the way in which these data can be used to estimate the dissociation constant of the interaction.
Topics: Animals; Autoradiography; Base Sequence; Cattle; DNA; DNA Footprinting; Deoxyribonuclease I; Electrophoresis; Isotope Labeling; Molecular Sequence Data; Plasmids; Polynucleotide 5'-Hydroxyl-Kinase; RNA-Directed DNA Polymerase
PubMed: 19997883
DOI: 10.1007/978-1-60327-418-0_10