Bi-functional cross-linking reagents efficiently capture protein-DNA complexes in Drosophila embryos Journal Article
| Authors: | Aoki, T.; Wolle, D.; Ben Noon, E. P.; Dai, Q.; Lai, E. C.; Schedl, P. |
| Article Title: | Bi-functional cross-linking reagents efficiently capture protein-DNA complexes in Drosophila embryos |
| Abstract: | Chromatin immunoprecipitation (ChIP) is widely used for mapping DNA-protein interactions across eukaryotic genomes in cells, tissues or even whole organisms. Critical to this procedure is the efficient cross-linking of chromatinassociated proteins to DNA sequences that are in close proximity. Since the mid-nineties formaldehyde fixation has been the method of choice. However, some protein-DNA complexes cannot be successfully captured for ChIP using formaldehyde. One such formaldehyde refractory complex is the developmentally regulated insulator factor, Elba. Here we describe a new embryo fixation procedure using the bi-functional cross-linking reagents DSG (disuccinimidyl glutarate) and DSP (dithiobis[succinimidyl propionate). We show that unlike standard formaldehyde fixation protocols, it is possible to capture Elba association with insulator elements in 2-5 h embryos using this new cross-linking procedure. We show that this new cross-linking procedure can also be applied to localize nuclear proteins that are amenable to ChIP using standard formaldehyde cross-linking protocols, and that in the cases tested the enrichment was generally superior to that achieved using formaldehyde cross-linking. © 2013 Landes Bioscience. |
| Keywords: | chromatin immunoprecipitation; dna binding; insensitive; chip; bi-functional crosslinkers; dsg dsp; elba; formadelhyde; insulators |
| Journal Title: | Fly |
| Volume: | 8 |
| Issue: | 1 |
| ISSN: | 1933-6934 |
| Publisher: | Taylor & Francis Group |
| Date Published: | 2014-10-17 |
| Start Page: | 44 |
| End Page: | 52 |
| Language: | English |
| DOI: | 10.4161/fly.26805 |
| PROVIDER: | scopus |
| PUBMED: | 24135698 |
| PMCID: | PMC3974894 |
| DOI/URL: | |
| Notes: | Export Date: 2 January 2014 -- Source: Scopus |
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