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| Status |
Public on Oct 20, 2020 |
| Title |
CUT&RUN reveals unique positioning of pre-initiated RNA polymerase II in the steady state of transcription |
| Organism |
Homo sapiens |
| Experiment type |
Genome binding/occupancy profiling by high throughput sequencing
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| Summary |
CUT&RUN is a powerful tool to study protein-DNA interactions in vivo. DNA fragments cleaved by the targeted micrococcal nuclease identify the footprints of DNA-binding proteins on the chromatin. We performed CUT&RUN on human lung carcinoma cell line A549 maintained in a multi-well cell culture plate to profile RNA polymerase II. Size-fractionation of aligned reads revealed that the collection of long (>270 bp) DNA fragments corresponded to the bimodal peak around the transcription start sites, as previously seen with chromatin immunoprecipitation. However, we found that shorter (<120 bp) fragments identify a well-defined peak localised at the transcription start sites. This distinct DNA footprint of shorter fragments, which constituted only about 5% of the total reads, indicates the unique positioning of the poised RNA polymerase II before transcription initiation, which has not been detected in the steady state of transcription by standard chromatin immunoprecipitation.
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| Overall design |
CUT&RUN assays targeting RNA polymerase II. Formaldehyde cell fixation was implemented in some of these assays.
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| Contributor(s) |
Miura M, Chen H |
| Citation(s) |
33070289 |
| |
| Submission date |
Aug 04, 2020 |
| Last update date |
Oct 24, 2020 |
| Contact name |
Michi Miura |
| E-mail(s) |
michi.miura.res@gmail.com
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| Organization name |
The University of Hong Kong
|
| Street address |
21 Sassoon Road
|
| City |
Hong Kong |
| ZIP/Postal code |
0000 |
| Country |
Hong Kong |
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| Platforms (1) |
| GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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| Samples (13)
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| Relations |
| BioProject |
PRJNA655178 |
| SRA |
SRP275783 |
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