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| Status |
Public on Sep 14, 2023 |
| Title |
Infertile patient blood sample 2 |
| Sample type |
SRA |
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| Source name |
Infertile patient blood
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| Organism |
Homo sapiens |
| Characteristics |
tissue: Infertile patient blood
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| Treatment protocol |
For tumor tissue, 0.5 cm3 tissue blocks were selected and minced through a 40 μm strainer to obtain single cell suspension. For blood samples, we directly took 1 ml of anticoagulated whole blood, and centrifuged at 1500 g/min for 10 min to collect blood cells.
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| Growth protocol |
K562 and COLO320-DM cells were grown in RPMI-1640 containing 10 % FBS；SW480 cells were grown in DMEM containing 10 % FBS
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were cross-linked with 0.5 % formaldehyde (Sigma) for 10 mins. The cross-linking reaction was terminated by glycine at a final concentration of 200 mM and lysed in lysis buffer (PBS contain 0.2% SDS) at room temperature for 5 min. After incubation, the nuclei were pelleted by centrifugation at 2,000 g/min for 5 min. The nuclei were transferred to 1.5 ml tubes and washed twice with PBS. For in situ restriction enzyme digestion, 140 μl of ddH2O, 20 μl of 10% Triton X-100, 20 μl of 10× NEBuffer 2.1, and 20 μl of HindIII (NEB, 20 units/μl) was added to the nuclei pellet and digested for 1.5 h at 37 °C in thermomixer (Eppendorf) with rotation at 1000 r.p.m. Add 5 μl of dNTP mix (10 mM each) and 5 μl of DNA polymerase I Klenow fragment (NEB, M0210) to the reaction system, place the sample in thermomixer with rotation at 37 °C at 1000 r.p.m for 30 mins. Add 27.5 μl of H2O, 3 μl of ATP (adenosine-triphosphate, 10mM), and 10 μl of T4 DNA ligase (Thermo, EL0011) to the reaction system, and placed the tube on the rotating mixers for 2 h at room temperature with rotation at 20 r.p.m. Add 20 μl of proteinase K (20 μg/ml) to the proximity ligation system, and then incubate at 60 °C for 2 hours. After digestion, the DNA was directly extracted using PCR Purification Kits (Zymo, D4013). DNA sequencing libraries were prepared using the VAHTS Universal Plus DNA Library Prep Kit (NDM627) according to manufacturer's protocol.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
DNBSEQ-T7 |
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| Data processing |
FastQC49 (version: 0.11.5) was used to assess the quality of raw reads, and FASTP50 (version: 0.23.2) was used to filter out the low-quality bases and adapter sequences. The clean read pairs which contained proximity ligation junction sequences “AAGCTAGCTT” were filtered out by Linux command line utility “grep”. The remaining reads were used for SNPs, indels, split reads, and CNV calling.
The remaining reads were aligned to the reference genome (hg19) and generated BAM file using BWA-MEM51 (version 0.7.17). The BAM file was sorted by SAMtools52 (version 1.15.1) and deduplicated by Sambamba53 (version 0.6.6). Next, we used BaseRecalibrator (GATK54, version 4.2.2) to calibrate the base quality scores, and HaplotypeCaller (GATK, version 4.2.2) to detect SNPs and indels.
The deduplicated BAM file generated in SNP and indel calling step were used to identify split reads. The split alignment reads were extracted by SAMtools (version 1.15.1) with command line “samtools view test_deduplicated.bam | grep SA > test_split_reads.txt”
BIC-seq255 was used to derive CNV segments from reads coverage data. For details, refer to the software manual “http://www.compbio.med.harvard.edu/BIC-seq/”. For the segmentation step, parameters were designed as binsize = 50,000 bp and λ = 2 to determine the final CNV breakpoints. chrY and chrM were excluded from the analysis.
The types and breakpoints of translocations were identified according to the split reads and CNV information. For unbalanced translocations, the chromosomal copy number at the breakpoint were usually altered, while balanced translocations do not have any chromosomal copy number changes
Assembly: hg19
Supplementary files format and content: .vcf for variations, .txt for copy number variation
Library strategy: MSV-Seq
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| Submission date |
Jun 01, 2022 |
| Last update date |
Sep 14, 2023 |
| Contact name |
zou yanyan |
| E-mail(s) |
[email protected]
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| Organization name |
HZAU
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| Street address |
SHIZISHAN
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| City |
WuHan |
| ZIP/Postal code |
430070 |
| Country |
China |
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| Platform ID |
GPL29480 |
| Series (2) |
| GSE205286 |
MGA-seq is an efficient tool for genetic variants and extrachromosomal DNA detection in cancer cell [MGA-Seq] |
| GSE205293 |
MGA-seq is an efficient tool for genetic variants and extrachromosomal DNA detection in cancer cell |
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| Relations |
| BioSample |
SAMN28814298 |
| SRA |
SRX15560471 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6209759_MSVSEQ_HC.hic |
169.5 Mb |
(ftp)(http) |
HIC |
| GSM6209759_MSVSEQ_HC_CNV.txt.gz |
98.9 Kb |
(ftp)(http) |
TXT |
| GSM6209759_MSVSEQ_HC_SV.vcf.gz |
1020.9 Kb |
(ftp)(http) |
VCF |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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