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| Status |
Public on Feb 24, 2021 |
| Title |
234_Wang_HiC_BWX4055_1mMIPTG_20min |
| Sample type |
SRA |
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| Source name |
Bacillus subtilis PY79
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| Organism |
Bacillus subtilis PY79 |
| Characteristics |
protocol: CH, 37degree C, 1mM IPTG for 20 min
genotype: parSdelta9 no a.b., -1degree parS no a.b., +26degree 12x XDS (loxP no a.b.), -19degree 12x XDS (loxP no a.b.), yvbJ::Pspank (optRBS) xerD (cat), deltaxerD (loxP-erm-loxP)
restriction enzyme: HindIII
strain: PY79
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| Treatment protocol |
Cells were growing in CH medium at 37˚C, IPTG was added to 1mM final concentration for 20min, fixed by 3% formaldehyde for 30min at room temperature
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| Growth protocol |
Bacillus subtilis strains were derived from the prototrophic strain PY79 (Youngman et al 1983, PMID: 6300908). Cells were grown in defined rich medium (CH) (Harwood and Cutting 1990, Molecular Biological Methods for Bacillus) at specified temperature. Staphylococcus aureus strains were derived from HG003 (Herbert et al 2010, PMID: 20212089). Cells were grown in tryptic soy broth (TSB) at specified temperature.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Hi-C experiments were performed using HindIII according to previous publications (Le et al, 2013, PMID: 24158908; Wang et al 2015, PMID: 26253537). ChIP-seq experiments were performed using anti-SMC, or anti-His antibodies according to previous publication (Graham et al 2014, PMID: 24829297).
Standard library construction protocol was used for Illumina HiSeq2500, Nextseq550, or MiSeq sequencing platforms. Illumina Truseq indexes were used.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
NextSeq 550 |
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| Description |
parS∆9 no a.b., -1 ̊ parS no a.b., +26˚ 12x XDS (loxP no a.b.), -19˚ 12x XDS (loxP no a.b.), yvbJ::Pspank (optRBS) xerD (cat), ∆xerD (loxP-erm-loxP)
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| Data processing |
Library strategy: HiC-Seq
Reads from each end of a DNA fragment were represented in each of the two FASTQ files generated by Illumina paired-end sequencing.
For Bacillus subtilis Hi-C, we mapped reads in each FASTQ file back to the genome of Bacillus subtilis PY79 (NCBI Reference Sequence: NC_022898.1) independently by Bowtie version 2.1.0 to preserve the order of reads. All the processing steps afterwards were performed using an in-house Perl scripts. For Staphylococcus aureus Hi-C, we mapped reads in each FASTQ file back to the genome of Staphylococcus aureus NCTC8325 genome (NCBI Reference Sequence: NC_007795.1) independently by Bowtie version 2.1.0 to preserve the order of reads. All the processing steps afterwards were performed using an in-house Perl scripts. For ChIP-seq, we mapped reads in each FASTQ file back to the genome of Bacillus subtilis PY79 (NCBI Reference Sequence: NC_022898.1) or Staphylococcus aureus NCTC8325 genome (NCBI Reference Sequence: NC_007795.1) independently using CLC genomics workbench 8.0, and exported .csv file.
For Hi-C, the two resulting SAM files were then merged together. Unaligned reads were discarded.
For Hi-C, the Bacillus subtilis (PY79) and Staphylococcus aureus (NCTC8325) genomes was divided into HindIII restriction fragments. Each aligned read was sorted into its corresponding restriction fragment. We inferred that a DNA fragment resulted from non-ligation or self-ligation if reads from both ends were from the same restriction fragment; these reads were discarded. Only DNA fragments for which the reads came from different restriction fragments were retained and used for construction of a Hi-C contact map.
For Hi-C, the genomes of Bacillus subtilis (PY79) and Staphylococcus aureus (NCTC8325) was then divided into bins 10 kb and the remaining reads were allocated to their corresponding bin. We then counted the number of fragments having reads with in different bins. A raw Hi-C contact map is the matrix of read counts in which each entry, mij, indicates the number of fragments with ends mapping to bins i and j. The raw Hi-C contact map/matrix is biased due to the uneven distribution of restriction enzyme sites and, to a lesser extent, differences in GC content and the mappability of individual reads. We normalized raw contact maps using an iterative normalization procedure (Imakaev et al., 2012 PMID: 22941365). Essentially, we converted the number of interactions, or read counts, into Hi-C scores by applying the following equation and iteratively repeating it for the resulting contact map after each cycle: mij =mij * (total reads) / (total reads in bin i * total reads in bin j). The iterative procedure was repeated until the maximum relative error of the total number of Hi-C scores in a bin was less than 10-5. Resulting matrices were normalized so that Hi-C scores for each row and column sum to 1. Subsequent analysis and visualization was done using R scripts.
Genome_build: NC_022898.1; NC_007795.1
Supplementary_files_format_and_content: Files ending with .matrix.txt: tab-delimited text files of the iteratively-corrected Hi-C contact maps/matrices. Files ending with .csv: comma-separated values files of ChIP-seq results.
Supplementary_files_format_and_content: matrix.txt files: 10kb bins of B. subtilis genome; normalized Hi-C interaction scores
Supplementary_files_format_and_content: csv files: genome position; reads; number of reads at each genome position
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| Submission date |
Feb 04, 2020 |
| Last update date |
Feb 25, 2021 |
| Contact name |
Xindan Wang |
| E-mail(s) |
xindan@iu.edu
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| Organization name |
Indiana University at Bloomington
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| Department |
Biology
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| Lab |
Biology Building 225
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| Street address |
1001 E 3rd St
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| City |
Bloomington |
| State/province |
IN |
| ZIP/Postal code |
47405 |
| Country |
USA |
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| Platform ID |
GPL28115 |
| Series (1) |
| GSE144742 |
XerD is required to unload bacterial SMC complexes at the replication terminus |
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| Relations |
| BioSample |
SAMN13981330 |
| SRA |
SRX7671102 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM4294720_234_Wang_HiC_BWX4055_1mMIPTG_20min.matrix.txt.gz |
152.4 Kb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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