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| Status |
Public on May 09, 2017 |
| Title |
Precise integration of inducible transcriptional elements (PrIITE) enables absolute control of inducible gene expression. |
| Organism |
Homo sapiens |
| Experiment type |
Expression profiling by high throughput sequencing
Genome binding/occupancy profiling by high throughput sequencing
|
| Summary |
Tetracycline-based inducible systems provide powerful methods for functional studies where gene expression can be controlled. However, the lack of tight control of the inducible system, leading to leakiness and adverse effects caused by undesirable tetracycline dosage requirements, has proven to be a limitation. Here we report that the combined use of genome editing tools and last generation Tet-On systems can resolve these issues. Our principle is based on precise integration of inducible transcriptional elements (coined PrIITE) targeted to; 1) exons of an endogenous gene of interest (GOI) and 2) a safe harbor locus. Using PrIITE cells harboring a GFP reporter or CDX2 transcription factor, we demonstrate discrete inducibility of gene expression with complete abrogation of leakiness. CDX2 PrIITE cells generated by this approach uncovered novel CDX2 downstream effector genes. Our results provide a strategy for characterization of dose-dependent effector functions of essential genes that require absence of endogenous gene expression.
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| Overall design |
Transcriptome and ChIP-seq analyses of inducible CDX2 integrated expression and wt CDX2 expression in LS174T colorectal cell lines by deep sequencing. In total six transcriptome data set (LS174T_wt; LS174T_CDX+/-; LS174T_CDX2-/-; LS174T_CDX2-/-1X induced CDX2 (4ng Dox); LS174T_CDX2-/-2X induced CDX2 (0ng Dox); LS174T_CDX2-/-2X induced CDX2; 4ng Dox); LS174T_CDX2-/-2X induced CDX2 (40ngDox)) and one ChIP-seq data set (ChIP-seq CDX2 ab)
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| Contributor(s) |
Bennet EP, Troelsen J, Hansen L |
| Citation(s) |
28472465, 33838073 |
| |
| Submission date |
Mar 31, 2017 |
| Last update date |
Jun 15, 2021 |
| Contact name |
Lars Hansen |
| E-mail(s) |
lah@sund.ku.dk
|
| Phone |
+45 35335499
|
| Organization name |
University of Copenhagen
|
| Department |
Department of Cellular and Molecular Medicine
|
| Lab |
Copenhagen Center for Glycomics (CCG)
|
| Street address |
3B Blegdamsvej
|
| City |
Copenhagen |
| ZIP/Postal code |
2200 N |
| Country |
Denmark |
| |
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| Platforms (1) |
| GPL17303 |
Ion Torrent Proton (Homo sapiens) |
|
| Samples (9)
|
| GSM2560482 |
LS174T CDX2 +/- clone 5B5 |
| GSM2560483 |
LS174T CDX2 -/- clone 3D6 |
| GSM2560484 |
LS174T CDX2-/- 1XindCDX2 (4 ng/ml Dox) clone 7D9 |
| GSM2560485 |
LS174T CDX2-/- 2XindCDX2 (0 ng/ml Dox) clone 6D6 |
| GSM2560486 |
LS174T CDX2-/- 2XindCDX2 (4 ng/ml Dox) clone 6D6 |
| GSM2560487 |
LS174T CDX2-/- 2XindCDX2 (40 ng/ml Dox) clone 6D6 |
|
| Relations |
| BioProject |
PRJNA381163 |
| SRA |
SRP102781 |
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