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Related Concept Videos

Chromatin Immunoprecipitation- ChIP02:36

Chromatin Immunoprecipitation- ChIP

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Chromatin immunoprecipitation, or ChIP, is an antibody-based technique used to identify sites on DNA that bind to transcription factors of interest or histone proteins. It also helps determine the type of histone modifications such as acetylation, phosphorylation, or methylation.
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Crossing over is the exchange of genetic information between homologous chromosomes during prophase I of meiosis I. Genetic recombination gives rise to allelic diversity in the newly formed daughter cells. In humans, crossing over produces genetically distinct haploid egg and sperm cells that undergo fertilization to produce unique offspring. Before cell division starts, the germ cell’s chromosome(s) undergo duplication in the S phase of the cell cycle. As the cells enter prophase I,...
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Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
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Immunoprecipitation, or IP, is a widely used technique that employs protein-antibody interactions to isolate proteins or protein complexes in their native state for studying protein-protein interactions, quaternary structures, or supramolecular complexes. Various modifications of the technique, including chromatin IP, cross-linking IP, and fluorescence IP, are commonly used.
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Efficient Chromatin Immunoprecipitation using Limiting Amounts of Biomass
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Mapping Recombination Initiation Sites Using Chromatin Immunoprecipitation.

Yan He1, Minghui Wang2, Qi Sun3

  • 1National Maize Improvement Center of China, Beijing Key Laboratory of Crop Genetic Improvement, China Agricultural University, Beijing, 100083, China.

Methods in Molecular Biology (Clifton, N.J.)
|August 12, 2016
PubMed
Summary

This study presents a chromatin immunoprecipitation (ChIP) protocol to map recombination initiation sites using RAD51 protein in maize. This method identifies double-strand break hotspots, aiding genome evolution research.

Keywords:
AntibodyChromatinChromosomesDouble-strand breaks (DSBs)ImmunoprecipitationMaizeRecombination

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Area of Science:

  • Genetics and Genomics
  • Molecular Biology

Background:

  • Genome-wide recombination maps offer insights into genome dynamics and evolution.
  • Understanding recombination initiation sites is crucial for studying genome stability.
  • Current methods may have limitations in precisely mapping these initiation sites.

Purpose of the Study:

  • To develop and describe a chromatin immunoprecipitation (ChIP) protocol for mapping recombination initiation sites in plants.
  • To utilize RAD51 protein as a marker for identifying double-strand break (DSB) formation hotspots.
  • To establish a method applicable across various species, including plants, animals, and fungi.

Main Methods:

  • Extraction of chromatin from meiotic flowers.
  • Shearing of chromatin and enrichment of fragments bound to RAD51 protein.
  • Identification of genomic locations of RAD51-bound fragments using next-generation sequencing.

Main Results:

  • A detailed chromatin immunoprecipitation (ChIP) protocol for mapping recombination initiation sites was successfully established using maize as a model.
  • The protocol effectively identifies double-strand break (DSB) formation hotspots by targeting the RAD51 protein.
  • The method demonstrated potential for broad applicability across different species.

Conclusions:

  • The described ChIP protocol provides a robust method for mapping meiotic recombination initiation sites.
  • This technique facilitates the study of genome dynamics, evolution, and DSB hotspots.
  • The protocol's adaptability makes it a valuable tool for diverse eukaryotic organisms.