2020
DOI: 10.1101/2020.08.10.243931
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Genome-wide analysis of DNA replication and DNA double strand breaks by TrAEL-seq

Abstract: Understanding the distribution of sites at which replication forks stall, and the ensuing fork processing events, requires genome-wide methods sensitive to both changes in replication fork structure and the formation of recombinogenic DNA ends. Here we describe Tr ansferase- A ctivated E nd L igation seq uencing (TrAEL-seq), a method that captures single stranded DNA 3’ ends genome-wide and with base pair resolution. TrAEL-seq labels DNA breaks, and profiles both stalled and processive replication fo… Show more

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Cited by 9 publications
(32 citation statements)
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“…We recently developed TrAEL-seq to detect replication fork stalling and replication intermediates (49). Unfortunately, TrAEL-seq is ineffective on copper-treated cells as copper-induced apoptotic fragments generate a high background (49,76) (example data is deposited at GEO GSE154811).…”
Section: Resultsmentioning
confidence: 99%
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“…We recently developed TrAEL-seq to detect replication fork stalling and replication intermediates (49). Unfortunately, TrAEL-seq is ineffective on copper-treated cells as copper-induced apoptotic fragments generate a high background (49,76) (example data is deposited at GEO GSE154811).…”
Section: Resultsmentioning
confidence: 99%
“…Plugs were equilibrated once in 100 μl 1x TdT buffer (NEB) for 30 min at room temperature, then incubated for 2 h at 37°C in 100 μl 1x TdT buffer containing 4 μl 10 mM ATP and 1 μl Terminal Transferase (NEB M0315L). Plugs were rinsed with 1 ml tris buffer (10 mM Tris HCl pH 8.0), equilibrated in 100 μl 1x T4 RNA ligase buffer (NEB) containing 40 μl 50% PEG 8000 for 1 hour at room temperature then incubated overnight at 25°C in 100 μl 1x T4 RNA ligase buffer (NEB) containing 40 μl 50% PEG 8000, 1 μl 10 pM/μl TrAEL-seq adaptor 1 (49) and 1 μl T4 RNA ligase 2 truncated KQ (NEB M0373L). Plugs were then rinsed with 1 ml tris buffer, transferred to 15 ml tubes and washed three times in 10 ml tris buffer with rocking at room temperature for 1-2 hours each, then washed again overnight under the same conditions.…”
Section: Methodsmentioning
confidence: 99%
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