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| Status |
Public on Jul 18, 2017 |
| Title |
Genome-wide maps of GlcNAc bound proteins and Pho ChIP-seq in Drosophila S2 cells and pupae. |
| Organism |
Drosophila melanogaster |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
We developed a strategy to map GlcNAc modified proteins based on a chemical chromatin precipitation strategy. Drosophila S2 cells, as well as wild type (wt) and sxc null pupae, were fed with 100 uM Ac4GalNAz and the resulting DNA was cross-linked, sonicated, and purified. DNA strands cross-linked to GlcNAz proteins were isolated by congugating with alkyn-biotin by Staudinger ligation followed by streptavidin purification. The resulting DNA was constructed into libraries for sequencing. To asses the robustness of our strategy, we compared GalNAz ChIP-seq results in S2 cells with two other GlcNAc ChIP-seq strategies, using a mutant β-1,4-galactosyltransferase (GalT) and the lecting wheat germ agglutinin (WGA). Briefly, GalT was incubated with cross-linked, sonicated and purified DNA along with UDP-GalNAz. Ligation to biotin with click chemistry followed by streptavidin purification resulted in library ready material. For WGA and Pho ChIP-seq, cross-linked and purified DNA was incubated with pho antibody or WGA resin and purified followed by library preperation.
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| Overall design |
Examination GlcNAc bound loci in Drosophila cells and pupae.
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| Contributor(s) |
Liu T, Myschyshyn M, Sinclair D, Cecioni S, Beja K, Honda BM, Morin RD, Vocadlo D |
| Citation(s) |
27918560 |
| Submission date |
Jul 18, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
David Vocadlo |
| E-mail(s) |
dvocadlo@sfu.ca
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| Organization name |
Simon Fraser University
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| Department |
Chemistry
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| Street address |
8888 University Drive
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| City |
Burnaby |
| State/province |
British Columbia |
| ZIP/Postal code |
V5A 1S6 |
| Country |
Canada |
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| Platforms (1) |
| GPL16479 |
Illumina MiSeq (Drosophila melanogaster) |
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| Samples (7)
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| GSM2238597 |
GlcNAc ChIP-seq on S2 cells using GalT purification |
| GSM2238598 |
input genomic DNA from S2 cells |
| GSM2238599 |
GlcNAc ChIP-seq on GalNAz fed wt pupae |
| GSM2238600 |
GlcNAc ChIP-seq on GalNAz fed sxc null pupae |
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| Relations |
| BioProject |
PRJNA329506 |
| SRA |
SRP078807 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE84502_RAW.tar |
277.1 Mb |
(http)(custom) |
TAR (of BEDGRAPH) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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