DNA Labeling Kit: A New Method for Targeted Labeling and Genomic Analysis of Nuclear Acids in Vivo
- 詳細技術說明
- Dr. Belmont from the University of Illinois at Urbana-Champaign has created a new system to determineDNA-Protein interactions. This method is more sensitive and take less time than the commonly usedmethods ChIP and DamID.
- *Abstract
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Dr. Belmont from the University of Illinois at Urbana-Champaign has created a new system to determineDNA-Protein interactions. This method is more sensitive and take less time than the commonly usedmethods ChIP and DamID.
How it works
This technology is a novel proximity-mapping approach capable ofidentifying DNA sequences that areorganized in or near any particularnuclear structure with adjustablespatial and temporal resolution. Thisapproach combines immunostainingand a tyramide-mediated labelingreaction, targets peroxidase enzyme to our interested nuclearcompartment and biotinylates nearbyDNA molecules, which can then beaffinity purified and identified. Thismethod can capture the changes ofchromosome domain organization in response to stimulation. Thisnew approach provides a powerfultool for understanding large-scalechromatin organization, chromatindynamics and gene positioning, aswell as their functional relationshipwith gene expression.
Advantages
- The inventors showed that tyramide radicals canlabel not only tyrosine moieties, but also DNA.Therefore, tyramide signal amplification can be usedto monitor DNA/DNA or DNA/Protein interactions.They can also attach biotin to the tyramide radical inorder to isolate DNA using streptavidin.
- This approach combines antibody staining with the Tyramide Signal Amplification (TSA) techniqueto label macromolecules surrounding a specifictarget. Using the TSA technique, the Horse RadishPeroxidase (HRP) is coupled with the antibody andbegins catalyzing the formation of tyramide-biotinfree radicals, which then diffuse short distances(prior to the covalent labeling of neighboringmacromolecules).
- The degree of labeling will be a function of distancefrom a given protein label after normalization for differences in accessibility and reactivity. Byantibody staining a specific protein concentrated ina particular compartment, this method can targetHRP to this protein and specifically label and identifyproteins and nucleic acids within this compartment.It also provides tools for studying chromatin domainorganization near specific compartments.
- 國家/地區
- 美國
