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| Status |
Public on Nov 23, 2022 |
| Title |
Azucena-flag_leaf-BS-seq-1 |
| Sample type |
SRA |
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| Source name |
flag_leaf
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| Organism |
Oryza sativa Japonica Group |
| Characteristics |
tissue: flag_leaf
chip antibody: n/a
genotype: wild type
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
For collection of meiotic spikelets with meiocytes at prophase of meiosis I, tillers whose flag leaf auricle was about 3 cm under the auricle of the penultimate leaf were chosen and 0.4-0.6 cm long fresh spikelets were collected. Meiotic spikelets collected above were put on a clean slide. The anthers were gently dissected out and collected using dissecting needles under the dissection microscope.The anthers were put on a clean slide with 5 µl DEPC water/1× PBS added. Under an inverted microscope (with 10× objective), a 1 ml syringe needle was use to dissect one end of the anthers. Because the meiocytes at this stage produce callose, they stick together and form worm-like cell clusters. Gentle pressure was applied to the anthers to allow the separation of wormlike cell clusters from anther walls. The cell clusters were collected with capillary glass pipettes without taking any somatic cells.
scBS-seq libraries were constructed using a reported protocol (Clark et al., 2017). The library was constructed using the CUT&Tag kit (Hyperactive® Universal CUT&Tag Assay Kit for Illumina, Vazyme Biotech Co.,Ltd).
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| Library strategy |
Bisulfite-Seq |
| Library source |
genomic |
| Library selection |
RANDOM |
| Instrument model |
Illumina NovaSeq 6000 |
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| Data processing |
Raw reads of all sequencing data were trimmed using fastp (v0.20.1)
The CUT&Tag clean reads were mapped to the rice IR64 reference genome (Qin P, Lu H, Du H, Wang H, Chen W, Chen Z, He Q, Ou S, Zhang H, Li X, Li X, Li Y, Liao Y, Gao Q, Tu B, Yuan H, Ma B, Wang Y, Qian Y, Fan S, Li W, Wang J, He M, Yin J, Li T, Jiang N, Chen X, Liang C, Li S. Pan-genome analysis of 33 genetically diverse rice accessions reveals hidden genomic variations. Cell. 2021 Jun 24;184(13):3542-3558.e16. doi: 10.1016/j.cell.2021.04.046. Epub 2021 May 28. PMID: 34051138.) and the rice Azucena reference genome (Zhou Y, Chebotarov D, Kudrna D, Llaca V, Lee S, Rajasekar S, Mohammed N, Al-Bader N, Sobel-Sorenson C, Parakkal P, Arbelaez LJ, Franco N, Alexandrov N, Hamilton NRS, Leung H, Mauleon R, Lorieux M, Zuccolo A, McNally K, Zhang J, Wing RA. A platinum standard pan-genome resource that represents the population structure of Asian rice. Sci Data. 2020 Apr 7;7(1):113. doi: 10.1038/s41597-020-0438-2. PMID: 32265447; PMCID: PMC7138821.) with Bowtie2 aligner (Langmead and Salzberg, 2012).
scBS-seq clean reads are "paired-end" data, but when matching, the F and R sides are used to match the reference genome separately using Bismark (v0.22.3) software to obtain bam files. The F-side and R-side bam files were then merged and the duplicate reads were removed, and the methylation site file was obtained using Bismark software.
Peaks were called uing MACS v2.1.0 with the significance cut-off q-value <=0.01
Duplications were removed, and uniquely mapped reads were retained for further analysis. Mean nuclear methylation levels were calculated by averaging methylation of individual cytosines in each context as described (Kim et al., 2019).
Supplementary files format and content: bigWig, narrowPeak,50bp/bin.map.gz,data analysis.xlsx
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| Submission date |
Nov 21, 2022 |
| Last update date |
Nov 23, 2022 |
| Contact name |
feng zhao |
| E-mail(s) |
[email protected]
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| Phone |
15926335800
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| Organization name |
Huazhong Agricultural University
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| Street address |
No. 1, Shizishan Street, Hongshan District
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| City |
Wuhan |
| State/province |
Hubei |
| ZIP/Postal code |
430072 |
| Country |
China |
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| Platform ID |
GPL27860 |
| Series (1) |
| GSE218416 |
Establish the male meiocyte-specific epigenomic landscape |
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| Relations |
| BioSample |
SAMN31813860 |
| SRA |
SRX18327110 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6744429_Azu_Flag_leaf_1_call_methylation.50bpbin.map.gz |
163.6 Mb |
(ftp)(http) |
MAP |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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