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| Status |
Public on Sep 15, 2022 |
| Title |
12-H-1 |
| Sample type |
SRA |
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| Source name |
Heart
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| Organism |
Mus musculus |
| Characteristics |
strain: C57BL/6J
tissue: Heart
age: 12 months
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| Treatment protocol |
Two million K562 or HeLa cells were seeded at 1 million cells per ml of culture medium without penicillin-streptomycin solution in 6-well plates. After 16 hr, cells were treated with 1.5 mM MMS (Sigma) for 1 hr or grown without MMS as control. One microgram of the K562 or HeLa genomic DNA was used directly as input into the SSiNGLe-AP protocol. For the MX treatment, after DNase fragmentation, the purified genomic DNA was incubated at 37C for 30 min in 20 µl of 50 mM K3PO4 (pH 7.0) buffer with or without 33 mM MX (Abmole). The DNA was recovered by ethanol precipitation, dissolved in 15 µl water and subjected to SSiNGLe-AP procedure starting from the blocking step preceded by denaturation. All samples were processed with SSiNGLe-AP protocol based on APE1, with exception of some samples where Endo IV was used instead as indicated.
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| Growth protocol |
Human CML leukemia cell line K562 was obtained from Cell Bank of Chinese Academy of Sciences, and human cervical carcinoma HeLa cell line was obtained from National Infrastructure of Cell Line Resource. Cells were cultured in RPMI 1640 (Thermo Fisher Scientific) supplemented with 10% (v/v) heat-inactivated fetal bovine serum (FBS, ExCell Bio) and 1% (v/v) pen-strep (HyClone) at 37 °C and 5% CO2.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
12 male mice of C57BL/6J strain from 4 age groups (3-, 12-, 19- and 22-month-old) were used to extract brains, hearts, livers, bone marrow, PBMCs and sperms. Red cells in bone marrow were removed by Red Blood Cell Lysis Buffer (Beyotime). PBMCs were isolated from blood using Ficoll-Paque PLUS (GE Healthcare). The motile spermatozoa were isolated from the mouse epididymal tail by collecting cells floating in the upper layer of RPMI 1640 (Thermo Fisher Scientific). The isolated tissues were immediately put into liquid nitrogen for quick freezing. The whole organs were cut into small pieces and ground into powder by tissue homogenizer in presence of liquid nitrogen to avoid heating. All ground tissue samples were stored at -80C prior to DNA isolation. Genomic DNA from human cell lines and the mouse brain, bone marrow, heart, liver and PBMCs was isolated using TIANamp Genomic DNA kit (Tiangen, DP304) with RNaseA treatment according to the manufacturer's protocols. Genomic DNA from sperm cells was extracted using Sperm DNA Purification Kit (Simgen, 4202050) following the manufacturer?s instructions. All DNA samples were eluted in UltraPure? DNase/RNase-Free Distilled Water (Invitrogen), measured the concentration using SMA6000 (Merinton) and stored at -20C.
Library construction method: SSiNGLe-AP method
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
The genomic and mitochondrial AP site mapping was performed the same way as in the SSiNGLe-ILM protocol (https://protocolexchange.researchsquare.com/article/pex-920/v2) with the two additional filtering steps. One, PCR duplicates were removed. Two, a read-pair was used only if the first base of the read 2 aligned to the reference sequence. The coordinate of one base upstream of the first aligning base of the read 2 was assigned as the position of the AP site. In the samples used for the chrM analysis, reads were aligned separately to the chrM genome sequences and processed in the same way. Then, the coordinates of AP sites mapping to chrM were added to the BED files containing AP sites mapping to the nuclear genome.
Genome_build: HG19(GRCh37) for human reads, mm10 (GRCm38) for the mouse reads and sequences corresponding to the bacterial spike-ins listed in the file spike-ins.fa for the reads from the spike-in control experiments.
Supplementary_files_format_and_content: The *spike-ins.bed files contain coordinates of AP sites in the sequences of spike-ins. All other BED files contain coordinates of AP sites mapped to either mouse or human genomes. The 4th columns show the number of reads representing the AP site.
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| Submission date |
Dec 15, 2021 |
| Last update date |
Sep 15, 2022 |
| Contact name |
Huifen Cao |
| E-mail(s) |
hfcao_bnu@163.com, hfcao@hqu.edu.cn
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| Phone |
+86 05926167250
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| Organization name |
Huaqiao University
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| Department |
School of Biomedical Sciences
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| Lab |
Institute of Genomics
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| Street address |
668 Jimei Road
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| City |
Xiamen |
| State/province |
Fujian |
| ZIP/Postal code |
361021 |
| Country |
China |
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| Platform ID |
GPL24247 |
| Series (1) |
| GSE190955 |
Complex Genomic Patterns of Abasic Sites in Mammalian DNA Revealed by High-Resolution SSiNGLe-AP Method |
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| Relations |
| BioSample |
SAMN24060807 |
| SRA |
SRX13439236 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM5736254_12-H-1.bed.gz |
6.5 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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