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| Status |
Public on Mar 20, 2024 |
| Title |
G4 quadruplex landscape and its regulation revealed by a new antibody capture method |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
Currently, the identification of G4 quadruplexes in vivo depends on a published and widely used G4-ChIP protocol (PMID: 29470465) that uses the anti-G4 antibody to captures the quadruplexes like regular chromatin. This method is challenged by some false positives and false negatives in G4-quadruplex capture. We have developed an antibody capture G4-ChIP (AbC G4-ChIP) method to overcome these challenges. The AbC G4-ChIP achieves (i) minimized in vitro artifacts during the incubation steps, and (ii) capture of G4-quadruplexes which are not protein-bound. We have applied the AbC G4-ChIP to study the regulatory effect of a CGG-repeat binding protein CGGBP1 on G4-quadruplex formation. The AbC G4-ChIP identifies inter-strand G/C-skew as a sequence property associated with CGGBP1-regulated G4-quadruplexes.
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| Overall design |
The antibody capture G4-ChIP (AbC G4-ChIP) employs permeabilization of fixed cells in presence of a buffer (Tris-KCl-Triton X100) such that 1H6 antibody could capture endogenous DNA G4-quadruplexes. The antibody-G4-quadruplex interaction is then crosslinked and pulled down. The AbC G4-ChIP ensures (i) minimization of the presence of in vitro G4-quadruplex artifacts (as false positives) formed during the assay, and (ii) loss of endogenous-formed G4-quadruplex (as false negatives) due to melting during crosslinking. The AbC G4-ChIP profile has been compared against both in vitro and conventional G4-ChIP profiles. The regulatory effect of CGG-repeat binding protein CGGBP1 has been studied in presence and absence of endogenous levels of CGGBP1. The functional relevance of genome-wide profile of CGGBP1-dependent G4-quadruplex regions has been analyzed.
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| Contributor(s) |
Datta S, Patel M, Patel D, Singh U |
| Citation(s) |
38484151 |
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| Submission date |
May 08, 2022 |
| Last update date |
Mar 21, 2024 |
| Contact name |
Umashankar Singh |
| Organization name |
IIT Gandhinagar
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| Department |
Biological Sciences and Engineering
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| Lab |
HoMeCell Lab
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| Street address |
Acad Block 4 Room 317 IIT Gandhinagar Palaj
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| City |
Gandhinagar |
| State/province |
Gujarat |
| ZIP/Postal code |
382055 |
| Country |
India |
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| Platforms (2) |
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| Samples (16)
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| GSM6122747 |
HEK293T cells, G4-ChIP |
| GSM6122748 |
HEK293T cells, G4-ChIP Input |
| GSM6122749 |
HEK293T cells, In vitro G4 DNA-IPnat |
| GSM6122750 |
HEK293T cells, In vitro G4 DNA-IPdenat |
| GSM6122751 |
HEK293T cells, In vitro Input |
| GSM6122752 |
HEK293T cells, CTut |
| GSM6122753 |
HEK293T cells, CTut Input |
| GSM6122754 |
HEK293T cells, KDut |
| GSM6122755 |
HEK293T cells, KDut Input |
| GSM6122756 |
HEK293T cells, CTtr |
| GSM6122757 |
HEK293T cells, CTtr Input |
| GSM6122758 |
HEK293T cells, KDtr |
| GSM6122759 |
HEK293T cells, KDtr Input |
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| Relations |
| BioProject |
PRJNA836187 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSE202456_RAW.tar |
970.0 Kb |
(http)(custom) |
TAR (of BED) |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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