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Sample GSM5227621 Query DataSets for GSM5227621
Status Public on Jul 26, 2022
Title WT_seedling_Hi-C
Sample type SRA
 
Source name WT seedling
Organism Arabidopsis thaliana
Characteristics ecotype: Col-0
genotype: WT
cell type: ten-day-old seedlings
library strategy: Hi-C
Growth protocol Plants were grown under long day (16 hr light; 8 hr dark) conditions at 22 °C.
Extracted molecule genomic DNA
Extraction protocol Ten-day-old seedlings were harvested and fixed with 20 ml 2% formaldehyde solution for 15 min in vacuum conditions at room temperature and then quenched by adding 2.162 ml 2.5 M glycine. Fixed seedling tissue was rinsed with water three times and dried with tissue paper. The nuclei were released by grinding in liquid nitrogen and then resuspended with 25 ml of extraction buffer I (0.4 M sucrose, 10 mM Tris HCl pH 8, 10 mM MgCl2, 5 mM β-mercaptoethanol, 0.1 mM PMSF, 13 μl protease inhibitor). Nuclei were filtered through miracloth (Calbiochem) and then centrifuged at 4000 rpm for 20 min at 4 °C. The supernatant was discarded whilst the pellet was resuspended with 1 ml of extraction buffer II (0.25 M sucrose, 10 mM Tris HCl pH 8, 10 mM MgCl2, 1% Triton X-100, 5mM β-mercaptoethanol, 0.1mM PMSF, 13 μl protease inhibitor). Then, the mixture was centrifuged at 14000 rpm for 10 min at 4 °C and the pellet was resuspended with 300 μl of extraction buffer III (1.7 M sucrose, 10 mM Tris HCl pH 8, 0.15% Triton X-100, 2 mM MgCl2, 5 mM β-mercaptoethanol, 0.1 mM PMSF, 1 μl protease inhibitor), load the mixture onto the top of an equal amount of clean extraction buffer III, then centrifuge at 14000 rpm for 10 min.
The Hi-C library construction and sequencing were conducted by Annoroad Gene Technology Co., Ltd (Beijing, China). Briefly, The pelleted nuclei were washed twice with 1x ice cold CutSmart buffer and finally resuspended in 0.5 ml volume. SDS was applied to permeabilize nuclei at 65 °C for 10 min, Triton X-100 was added to quench SDS. Thereafter, chromatin was digested with 400 units MboI overnight at 37 °C with gentle rocking. MboI was then denatured to cease activity. Digested chromatin underwent DNA end repair with biotin-14-dCTP insertion followed by blunt-end ligation. After decrosslinking with proteinase K at 65 °C, DNA was purified by phenol chloroform extraction method. Biotin-14-dCTP was removed from non-ligated DNA fragment ends using T4 DNA polymerase. DNA was sheared to a range of 200 to 600 bp by sonication. Next, the fragments underwent end repair and were pulled down by streptavidin C1 magnetic beads to enrich for fragments containing contact information. Fragment ends were then A-tailed, sequencing adapters were ligated, and libraries were amplified by PCR for 12-14 cycles. Following purification, libraries were sequenced using the Illumina HiSeq platform with 2×150 bp length.
 
Library strategy Hi-C
Library source genomic
Library selection other
Instrument model HiSeq X Ten
 
Data processing Hi-C: For Hi-C data assay, clean reads were mapped to TAIR10 genome using the HiC-Pro (version 2.11.1) pipeline. The bam files (bwt2merged.bam) generated by HiC-Pro containing mapping information were used as input files for Fan-C (version 0.9.8). The module ‘fanc auto’ was applied to generate 500 kb, 100 kb, 50 kb, 10 kb, 1kb contact matrix (hic files).
Genome_build: TAIR10
Supplementary_files_format_and_content: Hi-C: matrix and regions files of different resolution generated by Fan-C contain genomic contact and corresponding index information, respectively.
 
Submission date Apr 06, 2021
Last update date Jul 26, 2022
Contact name Xiaoqi Feng
E-mail(s) xiaoqi.feng@jic.ac.uk
Organization name John Innes Centre
Department Cell and Developmental Biology
Lab Xiaoqi Feng
Street address Norwich Research Park
City Norwich
State/province Norfolk
ZIP/Postal code NR4 7UH
Country United Kingdom
 
Platform ID GPL23157
Series (1)
GSE161366 Histone H2B.8 compacts flowering plant sperm through chromatin phase separation
Relations
BioSample SAMN18634027
SRA SRX10522767

Supplementary file Size Download File type/resource
GSM5227621_WT_seedling_100kb.matrix.gz 7.9 Mb (ftp)(http) MATRIX
GSM5227621_WT_seedling_100kb.regions.txt.gz 4.0 Kb (ftp)(http) TXT
GSM5227621_WT_seedling_10kb.matrix.gz 375.5 Mb (ftp)(http) MATRIX
GSM5227621_WT_seedling_10kb.regions.txt.gz 57.2 Kb (ftp)(http) TXT
GSM5227621_WT_seedling_500kb.matrix.gz 329.1 Kb (ftp)(http) MATRIX
GSM5227621_WT_seedling_500kb.regions.txt.gz 822 b (ftp)(http) TXT
GSM5227621_WT_seedling_rep1_1kb.matrix.txt.gz 1.3 Gb (ftp)(http) TXT
GSM5227621_WT_seedling_rep1_1kb.regions.txt.gz 578.3 Kb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file
Processed data are available on Series record

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