iDamIDseq and iDEAR

• Verfasst von: Gutiérrez Triana, José Arturo [VerfasserIn]
• Verfasst von: Mateo, Juan L. [VerfasserIn]
• Verfasst von: Ibberson, David [VerfasserIn]
• Verfasst von: Ryu, Soojin [VerfasserIn]
• Verfasst von: Wittbrodt, Joachim [VerfasserIn]
• Titel: iDamIDseq and iDEAR
• Titelzusatz: an improved method and computational pipeline to profile chromatin-binding proteins
• Verf.angabe: Jose Arturo Gutierrez-Triana, Juan L. Mateo, David Ibberson, Soojin Ryu and Joachim Wittbrodt
• E-Jahr: 2016
• Jahr: 6 September 2016
• Umfang: 7 S.
• Fussnoten: Gesehen am 28.08.2020
• Titel Quelle: Enthalten in: Development <Cambridge>
• Ort Quelle: Cambridge : The Company of Biologists, 1953
• Jahr Quelle: 2016
• Band/Heft Quelle: 143(2016), 22, Seite 4272-4278
• ISSN Quelle: 1477-9129
• DOI: doi:10.1242/dev.139261
• Datenträger: Online-Ressource
• Sprache: eng
• K10plus-PPN: 1727994205

Abstract

Skip to Next Section - DNA adenine methyltransferase identification (DamID) has emerged as an alternative method to profile protein-DNA interactions; however, critical issues limit its widespread applicability. Here, we present iDamIDseq, a protocol that improves specificity and sensitivity by inverting the steps DpnI-DpnII and adding steps that involve a phosphatase and exonuclease. To determine genome-wide protein-DNA interactions efficiently, we present the analysis tool iDEAR (iDamIDseq Enrichment Analysis with R). The combination of DamID and iDEAR permits the establishment of consistent profiles for transcription factors, even in transient assays, as we exemplify using the small teleost medaka (Oryzias latipes). We report that the bacterial Dam-coding sequence induces aberrant splicing when it is used with different promoters to drive tissue-specific expression. Here, we present an optimization of the sequence to avoid this problem. This and our other improvements will allow researchers to use DamID effectively in any organism, in a general or targeted manner.