A method for mapping intranuclear protein-DNA interactions and its application to a nuclease hypersensitive site.

Proceedings of the National Academy of Sciences of the United States of America ·Vol. 82 ·No. 8 ·1985-04-00 ·Pages 2296-300

Jackson PD, Felsenfeld G

Abstract

We have devised a method for mapping sites on DNA within the nucleus that are protected against nuclease attack by interaction with bound protein or other factors. This "footprinting" method uses an end-labeled sequence-specific DNA probe, which is annealed to the DNA from nuclear digests under carefully controlled conditions. The annealed complexes are treated with single-strand-specific nuclease, the resulting duplex molecules are electrophoresed on gels, and the gels are autoradiographed. The high sensitivity and resolution of the method have made it possible to obtain a detailed map of DNase I cutting patterns in the 5' flanking sequence of the chicken adult beta (beta A)-globin gene within nuclei from various tissues. In nuclei from adult erythrocytes, this domain is hypersensitive to nucleases. However, we detect within the domain two well-defined regions that are protected against attack, indicating the presence of one or more bound factors. Nuclei from oviduct or 5-day-old embryonic erythrocytes, in which the domain is not hypersensitive, show limited and different patterns of protection.

MeSH Terms

Animals Base Sequence Binding Sites Chick Embryo Chickens Chromosome Mapping DNA/genetics,metabolism Deoxyribonuclease I/metabolism Genes Globins/genetics Nucleoproteins/metabolism

Chemicals

Nucleoproteins Globins DNA Deoxyribonuclease I

Authors & Affiliations

2 authors, click to expand affiliations / ORCID

Jackson P D

Felsenfeld G

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24 references, click to expand

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Article Info

Journal

Proceedings of the National Academy of Sciences of the United States of America

Abbr.

Proc Natl Acad Sci U S A

ISSN

0027-8424

DOI

10.1073/pnas.82.8.2296

Published

1985-04-00

Pages

2296-300

Language

English

Region

United States

NLM ID

7505876

PMCID

PMC397544

Subset

External Links

PubMed source DOI: 10.1073/pnas.82.8.2296 PMC Full Text View journal details

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A method for mapping intranuclear protein-DNA interactions and its application to a nuclease hypersensitive site.

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