| Chapter title |
Immunoprecipitation of RNA:DNA Hybrids from Budding Yeast
|
|---|---|
| Chapter number | 8 |
| Book title |
DNA Topoisomerases
|
| Published in |
Methods in molecular biology, January 2018
|
| DOI | 10.1007/978-1-4939-7459-7_8 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-7458-0, 978-1-4939-7459-7
|
| Authors |
Aziz El Hage, David Tollervey |
| Abstract |
During transcription, the nascent transcript behind an elongating RNA polymerase (RNAP) can invade the DNA duplex and hybridize with the complementary DNA template strand, generating a three-stranded "R-loop" structure, composed of an RNA:DNA duplex and an unpaired non-template DNA strand. R-loops can be strongly associated with actively transcribed loci by all RNAPs including the mitochondrial RNA polymerase (mtRNAP). In this chapter, we describe two protocols for the detection of RNA:DNA hybrids in living budding yeast cells, one that uses conventional chromatin immunoprecipitation (ChIP-qPCR) and one that uses DNA:RNA immunoprecipitation (DRIP-qPCR). Both protocols make use of the S9.6 antibody, which is believed to recognize the intermediate A/B helical RNA:DNA duplex conformation, with no sequence specificity. |
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|---|---|---|
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Demographic breakdown
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|---|---|---|
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