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Article: Southwestern blotting in investigating transcriptional regulation

TitleSouthwestern blotting in investigating transcriptional regulation
Authors
Issue Date2008
PublisherNature Publishing Group. The Journal's web site is located at http://www.natureprotocols.com/about.php
Citation
Nature Protocols, 2008, v. 3 n. 1, p. 51-58 How to Cite?
AbstractSouthwestern blotting is used to investigate DNA-protein interactions. The advantage of this technique over other related methods such as electrophoretic mobility shift assay (EMSA) and DNA footprinting is that it provides information regarding the molecular weight of unknown protein factor. This method combines the features of Southern and Western blotting techniques; a denaturing SDS-PAGE is first employed to separate proteins electrophoretically based on size, and after transferring the proteins to a membrane support, the membrane-bound proteins are renatured and incubated with a 32P-labeled double-stranded oligonucleotide probe of specific DNA sequence. The interaction of the probe with the protein(s) is later visualized by autoradiography. This technique could be combined with database searching (TransFac, http://www.gene-regulation.com/pub/ databases.html#transfac), prediction of potential protein factors binding onto a target motif (e.g., Patch search), in vitro supershift EMSA and in vivo chromatin immunoprecipitation (ChIP) assays for effective identification of protein factors. The whole Southwestern blotting procedure takes ∼4 d to complete. In this article, a commonly used protocol and expected results are described and discussed.
Persistent Identifierhttp://hdl.handle.net/10722/89291
ISSN
2015 Impact Factor: 9.646
2015 SCImago Journal Rankings: 9.039
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorSiu, FKYen_HK
dc.contributor.authorLee, LTOen_HK
dc.contributor.authorChow, BKCen_HK
dc.date.accessioned2010-09-06T09:55:00Z-
dc.date.available2010-09-06T09:55:00Z-
dc.date.issued2008en_HK
dc.identifier.citationNature Protocols, 2008, v. 3 n. 1, p. 51-58en_HK
dc.identifier.issn1754-2189en_HK
dc.identifier.urihttp://hdl.handle.net/10722/89291-
dc.description.abstractSouthwestern blotting is used to investigate DNA-protein interactions. The advantage of this technique over other related methods such as electrophoretic mobility shift assay (EMSA) and DNA footprinting is that it provides information regarding the molecular weight of unknown protein factor. This method combines the features of Southern and Western blotting techniques; a denaturing SDS-PAGE is first employed to separate proteins electrophoretically based on size, and after transferring the proteins to a membrane support, the membrane-bound proteins are renatured and incubated with a 32P-labeled double-stranded oligonucleotide probe of specific DNA sequence. The interaction of the probe with the protein(s) is later visualized by autoradiography. This technique could be combined with database searching (TransFac, http://www.gene-regulation.com/pub/ databases.html#transfac), prediction of potential protein factors binding onto a target motif (e.g., Patch search), in vitro supershift EMSA and in vivo chromatin immunoprecipitation (ChIP) assays for effective identification of protein factors. The whole Southwestern blotting procedure takes ∼4 d to complete. In this article, a commonly used protocol and expected results are described and discussed.en_HK
dc.languageengen_HK
dc.publisherNature Publishing Group. The Journal's web site is located at http://www.natureprotocols.com/about.phpen_HK
dc.relation.ispartofNature Protocolsen_HK
dc.titleSouthwestern blotting in investigating transcriptional regulationen_HK
dc.typeArticleen_HK
dc.identifier.openurlhttp://library.hku.hk:4550/resserv?sid=HKU:IR&issn=1750-2799&volume=3&spage=51&epage=58&date=2008&atitle=Southwestern+blotting+in+investigating+transcriptional+regulationen_HK
dc.identifier.emailLee, LTO: ltolee2@hkucc.hku.hken_HK
dc.identifier.emailChow, BKC: bkcc@hku.hken_HK
dc.identifier.authorityLee, LTO=rp00727en_HK
dc.identifier.authorityChow, BKC=rp00681en_HK
dc.description.naturelink_to_subscribed_fulltext-
dc.identifier.doi10.1038/nprot.2007.492en_HK
dc.identifier.pmid18193021-
dc.identifier.scopuseid_2-s2.0-38149003098en_HK
dc.identifier.hkuros140748en_HK
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-38149003098&selection=ref&src=s&origin=recordpageen_HK
dc.identifier.volume3en_HK
dc.identifier.issue1en_HK
dc.identifier.spage51en_HK
dc.identifier.epage58en_HK
dc.identifier.eissn1750-2799-
dc.identifier.isiWOS:000253140400006-
dc.identifier.scopusauthoridSiu, FKY=6701518484en_HK
dc.identifier.scopusauthoridLee, LTO=8367269000en_HK
dc.identifier.scopusauthoridChow, BKC=7102826193en_HK
dc.identifier.citeulike2761685-

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