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| Status |
Public on Apr 16, 2015 |
| Title |
Identification of RNAs associated with telomeres by enChIP-RNA-Seq |
| Organism |
Mus musculus |
| Experiment type |
Expression profiling by high throughput sequencing
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| Summary |
Accumulating evidence suggests important roles of RNAs interacting with genomic regions in the regulation of genome functions including X chromosome inactivation and gene expression. However, no method to identify RNAs interacting with a given genomic region in a non-biased manner has been reported. Here, we used engineered DNA-binding molecule-mediated chromatin immunoprecipitation (enChIP) combined with the RNA-Seq analysis (enChIP-RNA-Seq) to perform non-biased search of RNAs interacting with telomeres. In enChIP-RNA-Seq, the target genomic regions are captured with an engineered DNA-binding molecule such as a TAL protein. Subsequently, RNAs are purified and subjected to the RNA-Seq analysis. The detected RNAs contained known telomere-binding RNAs including telomerase RNA and Cajal body-specific RNAs. In addition, we detected many novel telomere-binding RNAs. We confirmed binding of candidate RNAs to telomeres by the enChIP-RT-PCR analysis. Identified novel telomere-binding RNAs may play important roles in telomere functions. In addition, our results suggest that enChIP-RNA-Seq analysis would be useful for identification of RNAs interacting with specific genomic regions.
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| Overall design |
RNAs associated with telomeres were identified by using the enChIP technology combined with deep sequencing using Illumina Miseq. Briefly, to isolate telomeres, a TAL protein, Telomere-TAL (Tel-TAL), recognizing a 19-bp sequence containing an array of TTAGGG (telomere repeats) was fused with 3xFLAG tag and NLS (3xFN-Tel-TAL) and LexA protein (3xFNLDD)11 were expressed in a mouse hematopoietic cell line, Ba/F3, respectively. The cells were crosslinked with formaldehyde, and crosslinked chromatin was fragmented by sonication. Subsequently, chromatin complexes containing 3xFN-Tel-TAL or 3xFNLDD were immunoprepicitated with anti-FLAG M2 Ab.
Supplementary URL: http://www.nature.com/srep/2013/131108/srep03171/full/srep03171.html
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| Contributor(s) |
Okuzaki D, Fujita T, Fujii H |
| Citation(s) |
25874893 |
| BioProject |
PRJNA257682 |
| Submission date |
Aug 14, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Daisuke Okuzaki |
| E-mail(s) |
dokuzaki@biken.osaka-u.ac.jp
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| Phone |
+81-6-6879-4935
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| Organization name |
Osaka univ.
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| Department |
Immunology Frontier Research Center
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| Lab |
Human Immunology （Single Cell Genomics)
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| Street address |
Yamadaoka 3-1
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| City |
Suita |
| State/province |
Osaka |
| ZIP/Postal code |
565-0871 |
| Country |
Japan |
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| Platforms (1) |
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| Samples (2) |
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| Relations |
| SRA |
SRP045319 |
