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Article: Incorporating the ABI GeneScan® Analysis to a RACE-based technique for mapping multiple transcription initiation sites

TitleIncorporating the ABI GeneScan® Analysis to a RACE-based technique for mapping multiple transcription initiation sites
Authors
Issue Date2001
Citation
Applied Biochemistry And Biotechnology - Part B Molecular Biotechnology, 2001, v. 17 n. 2, p. 129-134 How to Cite?
AbstractDetermination of transcription initiation sites has commonly been performed by primer extension and RNase protection assay using radioactively labeled oligonucleotides. Recently, a protocol based on modified 5′ rapid amplification of cDNA ends (RACE) with the use of fluorescently labeled primer was developed. Here, we describe the use of RACE-based technique in conjunction with the GeneScan® analysis for the determination of transcription initiation sites of genes of interest. The RACE technique is based on the ligation of an adapter to both ends of the cDNAs. The gene of interest was first amplified by PCR using a gene-specific and a 5′ adapter primer. Subsequently, nested PCR was performed using an internal gene-specific primer paired with a fluorescently end-labeled adapter primer. The size of the fluorescently labeled PCR products was directly determined by the ABI PRISM 377 GeneScan® Analyzer. This novel approach provides an accurate, sensitive, and convenient method for mapping transcription initiation sites, especially for genes with multiple transcription initiation sites, for genes expressed at low levels, and for splice variants that display alternative splicing farther than a few hundred nucleotides downstream from the transcription initiation site. This article describes the application of this new method in the mapping of transcription initiation sites of two splice variants of rat frizzled related protein transcripts.
Persistent Identifierhttp://hdl.handle.net/10722/148228
ISSN
2015 Impact Factor: 1.752
2015 SCImago Journal Rankings: 0.787
ISI Accession Number ID
References

 

DC FieldValueLanguage
dc.contributor.authorPing Yam, JWen_US
dc.contributor.authorKoon, WCen_US
dc.contributor.authorWendy Hsiao, WLen_US
dc.date.accessioned2012-05-29T06:11:38Z-
dc.date.available2012-05-29T06:11:38Z-
dc.date.issued2001en_US
dc.identifier.citationApplied Biochemistry And Biotechnology - Part B Molecular Biotechnology, 2001, v. 17 n. 2, p. 129-134en_US
dc.identifier.issn1073-6085en_US
dc.identifier.urihttp://hdl.handle.net/10722/148228-
dc.description.abstractDetermination of transcription initiation sites has commonly been performed by primer extension and RNase protection assay using radioactively labeled oligonucleotides. Recently, a protocol based on modified 5′ rapid amplification of cDNA ends (RACE) with the use of fluorescently labeled primer was developed. Here, we describe the use of RACE-based technique in conjunction with the GeneScan® analysis for the determination of transcription initiation sites of genes of interest. The RACE technique is based on the ligation of an adapter to both ends of the cDNAs. The gene of interest was first amplified by PCR using a gene-specific and a 5′ adapter primer. Subsequently, nested PCR was performed using an internal gene-specific primer paired with a fluorescently end-labeled adapter primer. The size of the fluorescently labeled PCR products was directly determined by the ABI PRISM 377 GeneScan® Analyzer. This novel approach provides an accurate, sensitive, and convenient method for mapping transcription initiation sites, especially for genes with multiple transcription initiation sites, for genes expressed at low levels, and for splice variants that display alternative splicing farther than a few hundred nucleotides downstream from the transcription initiation site. This article describes the application of this new method in the mapping of transcription initiation sites of two splice variants of rat frizzled related protein transcripts.en_US
dc.languageengen_US
dc.relation.ispartofApplied Biochemistry and Biotechnology - Part B Molecular Biotechnologyen_US
dc.subject.meshAlternative Splicingen_US
dc.subject.meshAnimalsen_US
dc.subject.meshBase Sequenceen_US
dc.subject.meshChromosome Mapping - Methods - Statistics & Numerical Dataen_US
dc.subject.meshDna Primers - Geneticsen_US
dc.subject.meshGenetic Techniques - Statistics & Numerical Dataen_US
dc.subject.meshGenetic Variationen_US
dc.subject.meshProteins - Geneticsen_US
dc.subject.meshRandom Amplified Polymorphic Dna Technique - Methods - Statistics & Numerical Dataen_US
dc.subject.meshRatsen_US
dc.subject.meshRibonucleasesen_US
dc.subject.meshTranscription, Geneticen_US
dc.titleIncorporating the ABI GeneScan® Analysis to a RACE-based technique for mapping multiple transcription initiation sitesen_US
dc.typeArticleen_US
dc.identifier.emailPing Yam, JW:judyyam@pathology.hku.hken_US
dc.identifier.authorityPing Yam, JW=rp00468en_US
dc.description.naturelink_to_subscribed_fulltexten_US
dc.identifier.doi10.1385/MB:17:2:129en_US
dc.identifier.pmid11395861en_US
dc.identifier.scopuseid_2-s2.0-0034996250en_US
dc.identifier.hkuros57558-
dc.relation.referenceshttp://www.scopus.com/mlt/select.url?eid=2-s2.0-0034996250&selection=ref&src=s&origin=recordpageen_US
dc.identifier.volume17en_US
dc.identifier.issue2en_US
dc.identifier.spage129en_US
dc.identifier.epage134en_US
dc.identifier.isiWOS:000168804500004-
dc.publisher.placeUnited Statesen_US

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