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    Protein Tagging for Chromatin Immunoprecipitation from Arabidopsis
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    Keywords:
    Chromatin immunoprecipitation
    Immunoprecipitation
    ChIP-sequencing
    Chromatin immunoprecipitation
    Immunoprecipitation
    ChIP-on-chip
    ChIP-sequencing
    Methylated DNA immunoprecipitation
    Citations (0)
    Chromatin immunoprecipitation (ChIP) is a widely used method to explore in vivo interactions between proteins and DNA. The ChIP assay takes several days to complete, involves several tube transfers and uses either phenol–chlorophorm or spin columns to purify DNA. The traditional ChIP method becomes a challenge when handling multiple samples. We have developed an efficient and rapid Chelex resin-based ChIP procedure that dramatically reduces time of the assay and uses only a single tube to isolate PCR-ready DNA. This method greatly facilitates the probing of chromatin changes over many time points with several antibodies in one experiment.
    Chromatin immunoprecipitation
    Immunoprecipitation
    ChIP-sequencing
    ChIP-on-chip
    Citations (225)
    Sequential chromatin immunoprecipitation (ChIP) is commonly used to investigate DNA-protein and protein-protein interactions to a specific genomic region. However, it can be tricky to achieve a robust and reproducible signal with sequential ChIP. Here, we provide an optimized two-step ChIP protocol to quantify the in vivo associates of multiple proteins with the same DNA regulatory element. For complete details on the use and execution of this protocol, please refer to He et al. (2020).
    Chromatin immunoprecipitation
    Immunoprecipitation
    ChIP-sequencing
    ChIP-on-chip
    Immunoprecipitation
    Chromatin immunoprecipitation
    ChIP-on-chip
    ChIP-sequencing
    genomic DNA
    Chromatin-immunoprecipitation (ChIP) is a powerful technique for mapping the protein-DNA interactions that occur in living cells. The critical technical determinant for successful ChIP is the availability of an appropriate, "ChIP-grade" serum. Here we present a technique designed to assess whether sera are suitable for ChIP, and to quantify their efficiency relative to a positive internal reference. This approach is useful as a first step toward ChIP-on-chip or ChIP-sequencing, especially in the case of recently identified proteins for which no binding sites are yet known.
    Chromatin immunoprecipitation
    Immunoprecipitation
    ChIP-on-chip
    ChIP-sequencing
    Citations (5)
    Chromatin immunoprecipitation (ChIP) is a technique widely used for determining the genomic location of modified histones and other chromatin-associated factors. Here we describe the methodology we have used in our laboratory for the immunoprecipitation of chromatin isolated from cells in the absence of crosslinking. Chromatin released from nuclei by micrococcal nuclease digestion is centrifuged through sucrose gradients to allow selection of monoor dinucleosomes. This allows a protein or modification at a particular gene or locus to be mapped at higher resolution than in a crosslinked ChIP experiment. Two methods for the immunoprecipitation of chromatin are described: a large-scale fractionation by which it is possible to visualize the proteins of the immunoprecipitate by polyacrylamide gel electrophoresis, PAGE and a small-scale method that is more appropriate when the quantity of chromatin is limited. The sequence content of DNA extracted from the immunoprecipitated chromatin is analyzed by hybridization of Southern or slot blots, or by quantitative polymerase chain reaction. Enrichment of particular sequences in the immunoprecipitated fraction reveals the presence and extent of the modification at this location.
    Chromatin immunoprecipitation
    Immunoprecipitation
    Micrococcal nuclease
    ChIP-sequencing
    ChIP-on-chip
    Nuclease
    ChIA-PET
    Citations (35)