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Sample GSM5736279 Query DataSets for GSM5736279
Status Public on Sep 15, 2022
Title 12-M-2
Sample type SRA
 
Source name Bone marrow
Organism Mus musculus
Characteristics strain: C57BL/6J
tissue: Bone marrow
age: 12 months
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.
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.
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
 
Library strategy OTHER
Library source genomic
Library selection other
Instrument model Illumina NovaSeq 6000
 
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.
 
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
Phone +86 05926167250
Organization name Huaqiao University
Department School of Biomedical Sciences
Lab Institute of Genomics
Street address 668 Jimei Road
City Xiamen
State/province Fujian
ZIP/Postal code 361021
Country China
 
Platform ID GPL24247
Series (1)
GSE190955 Complex Genomic Patterns of Abasic Sites in Mammalian DNA Revealed by High-Resolution SSiNGLe-AP Method
Relations
BioSample SAMN24060777
SRA SRX13439213

Supplementary file Size Download File type/resource
GSM5736279_12-M-2.bed.gz 7.0 Mb (ftp)(http) BED
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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