General Information
- Experiment ID
- OEX002081
- Experiment Name
- NAIL-seq_EdU_HU
- Experiment Type
- Genomic
- Description
- NAIL-seq
- Protocol
- Cells with indicated treatment were harvested, washed with PBS, and applied to genomic DNA extraction with Proteinase K digestion as described previously (Yin et al., 2019). Purified genomic DNA was subjected to Click reaction with 100 µM Biotin-Azide, 2 mM CuSO4, 4 mM THPTA, and 10 mM Sodium ascorbate at 25℃ for 1 hour, followed by DNA precipitation. Dissolved DNA was denatured at 95℃ for 5 min and phosphorylated by T4 PNK at 37℃ for 30 min. For EdU- and/(or) BrdU-labelled samples, the denatured and modified DNA were purified by anti-BrdU (BU1/75) and Protein G beads for BrdU-labeled proportion, and the supernatants after anti-BrdU-ChIP were denatured again and then incubated with Streptavidin C1 beads (Dynabeads) to isolate EdU-labeled proportion. For EdU/HU labeled samples, T4 PNK-treated DNA were incubated with C1 beads directly. All the enrichment was incubated at room temperature for 4 hours. Enriched EdU- or BrdU-labeled ssDNA were ligated on-beads with two types of bridge adapters (Bridge adapter-1 and -2, see key resource table for sequence details) at room temperature for at least 4 hours as follow: 1 µM Bridge adapters each, 1,600 U T4 DNA ligase and 15% PEG-8000. After that, ligated DNA were harshly washed and tagged with Illumina P5-I5 and P7-I7 sequence as described (Yin et al., 2019). All the libraries were subjected to 2 × 150 bp Hiseq sequencing.
Experiment information
- Attributes
-
library_strategy Targeted-Capture library_selection ChIP library_layout Paired mate_pair Y platform Illumina HiSeq 2000 - Project
-
1
Project ID Project name Create Date OEP000658 Transcription orchestrates early DNA replication initiation in mammalian cells 2019-11-05 - Samples
-
8
Sample ID Sample Name Organism Tissue Create Date OES090599 mESC_Nailseq_EH_WT-IAA Mus musculus cell line 2021-05-08 OES090597 mESC_Nailseq_EH_Pol2-IAA Mus musculus cell line 2021-05-08 OES090598 mESC_Nailseq_EH_Pol2 Mus musculus cell line 2021-05-08 OES090600 mESC_Nailseq_EH_WT Mus musculus cell line 2021-05-08 OES090601 mouseSB_Nailseq_EH_ama0 Mus musculus Splenic B cells 2021-05-08 OES090602 mouseSB_Nailseq_EH_ama0.5 Mus musculus Splenic B cells 2021-05-08 OES090603 mouseSB_Nailseq_EH_ama1 Mus musculus Splenic B cells 2021-05-08 OES090604 mouseSB_Nailseq_EH_ama2 Mus musculus Splenic B cells 2021-05-08 - Runs
-
8
Run ID Run Name Sample Data Num Create Date OER181041 mESC_Nailseq_EH_WT-IAA_HU059_LY138b mESC_Nailseq_EH_WT-IAA 2 2021-05-08 OER181042 mESC_Nailseq_EH_Pol2-IAA_HU059_LY138d mESC_Nailseq_EH_Pol2-IAA 2 2021-05-08 OER181043 mESC_Nailseq_EH_Pol2_HU059_LY138c mESC_Nailseq_EH_Pol2 2 2021-05-08 OER181044 mESC_Nailseq_EH_WT_HU037_LY096h mESC_Nailseq_EH_WT 2 2021-05-08 OER181045 mouseSB_Nailseq_EH_ama0_HU020_LY069b mouseSB_Nailseq_EH_ama0 2 2021-05-08 OER181046 mouseSB_Nailseq_EH_ama0.5_HU020_LY069c mouseSB_Nailseq_EH_ama0.5 2 2021-05-08 OER181047 mouseSB_Nailseq_EH_ama1_HU020_LY069d mouseSB_Nailseq_EH_ama1 2 2021-05-08 OER181048 mouseSB_Nailseq_EH_ama2_HU020_LY069e mouseSB_Nailseq_EH_ama2 2 2021-05-08
Author Information
- Create Date
- 2019-11-05
- Last Modified
- 2019-11-07
- Submission
- Chen Ai