期刊
NUCLEIC ACIDS RESEARCH
卷 41, 期 5, 页码 -出版社
OXFORD UNIV PRESS
DOI: 10.1093/nar/gks1342
关键词
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资金
- Fonds voor Wetenschappelijk Onderzoek-Vlaanderen
- Vlaams Interuniversiair Instituut (VIB)
- Onderzoeksfonds (OZR-GOA), Vrije Universiteit Brussel
Nanobodies (R) are single-domain antibody fragments derived from camelid heavy-chain antibodies. Because of their small size, straightforward production in Escherichia coli, easy tailoring, high affinity, specificity, stability and solubility, nanobodies (R) have been exploited in various biotechnological applications. A major challenge in the post-genomics and post-proteomics era is the identification of regulatory networks involving nucleic acid-protein and protein-protein interactions. Here, we apply a nanobody (R) in chromatin immunoprecipitation followed by DNA microarray hybridization (ChIP-chip) for genome-wide identification of DNA-protein interactions. The Lrp-like regulator Ss-LrpB, arguably one of the best-studied specific transcription factors of the hyperthermophilic archaeon Sulfolobus solfataricus, was chosen for this proof-of-principle nanobody (R)-assisted ChIP. Three distinct Ss-LrpB-specific nanobodies (R), each interacting with a different epitope, were generated for ChIP. Genome-wide ChIP-chip with one of these nanobodies (R) identified the well-established Ss-LrpB binding sites and revealed several unknown target sequences. Furthermore, these ChIP-chip profiles revealed auxiliary operator sites in the open reading frame of Ss-lrpB. Our work introduces nanobodies (R) as a novel class of affinity reagents for ChIP. Taking into account the unique characteristics of nanobodies (R), in particular, their short generation time, nanobody (R)-based ChIP is expected to further streamline ChIP-chip and ChIP-Seq experiments, especially in organisms with no (or limited) possibility of genetic manipulation.
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