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| Status |
Public on Jun 13, 2016 |
| Title |
osdrm2 smRNA seq |
| Sample type |
SRA |
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| Source name |
DongJin
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| Organism |
Oryza sativa Japonica Group |
| Characteristics |
tissue: 12 day seedling
mutant: osdrm2
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| Growth protocol |
ere germinated and grown on the hormone-free, half-strength Murashige & Skoog (MS) medium under 16/8 hrs light/dark at 30/25°C,Twelve-day-old seedling leaves from each genotype were harvested for chromatin, genomic DNA or total RNA extraction
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| Extracted molecule |
total RNA |
| Extraction protocol |
Rice seedling total RNA samples were isolated using TRIzol reagent (Invitrogen).Seedling were crosslinked in 1% formaldehyde under vacuum. Chromatin was extracted and fragmented to 200-750 bp by sonication, and ChIP was performed using the following antibodies: H3K27me3 (a2363, AbclonalABclonal ).The eluted ChIP DNA was used to generate Illumina sequencing libraries following the manufacturer's protocol.
RNA libraries were prepared according to the protocol strictly (http://support.illumina.com/content/dam/illumina-support/documents/documentation/chemistry_documentation/samplepreps_truseq/truseqstrandedmrna/truseq-stranded-mrna-sample-prep-guide-15031047-e.pdf). Briefly, 1-4 μg of total RNAs were used for mRNA purification and cDNA synthesis. After single ‘A’ nucleotide was added to the 3’ ends of the blunt fragments of the double strand cDNAs, indexing adapters are ligated to the ends of the cDNA. PCR are performed to amplify the DNA fragments. The amplified DNA fragments were purified and sequenced with the Illumina Hiseq-2000 system.The eluted ChIP DNA was used to generate Illumina sequencing libraries following the manufacturer's protocol.
RNA-Seq
ChIP-Seq
Bs-seq
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| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 2000 |
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| Data processing |
Quality Control: fastqc
Remove low quality data:BS-seq raw data, ChIP-seq raw data and RNA-seq raw data's low quality reads were removed from raw data by Trimmomatic. smRNA sequence raw data were cleaned by removing adaptor contamination and low qualityReads by CutAdaptor (Martin, 2011). miRNA (http://www.mirbase.org/ftp.shtml), tRNA(http://rice.plantbiology.msu.edu/analyses_search_tRNA.shtml), rRNA (Rice GenomeAnnotation MSU7.0), snoRNA and snRNA (http://www.arb-silva.de/no_cache/download/archive/release_111/Exports/) were also removed.
Mapping: BS-seq clean data were mapped to MSU7.0 rice reference genome (ftp://ftp.plantbiology.msu.edu/pub/data/Eukaryotic_Projects/o_sativa/annotation_dbs/pseudomolecules/version_7.0/all.dir/) by BatMeth (Lim et al., 2012).The RNA-seq clean data were mapped against the MSU7.0 rice genome with corresponding annotation by Tophat2.0(Kim et al., 2013) (http://tophat.cbcb.umd.edu/) using default parameters.And then, the genes’ count were calculated with htseq-count, and the differentially expression’s genes (|log (FC)|>=2 && qvalue < 0.05) were calculated by DESeq2.ChIP-seq Clean reads were aligned to the rice MSU7.0 reference genome by Bowtie (Langmead et al., 2009) allowing 0 mismatches and unique mapping.The ChIP-seq clean reads were mapped to the rice genome (RGAP version 7) by default allowing up to 2 mismatch using Bowtie2 (version 2.1.0).Only uniquely mapped reads were retained for further analysis.
Two types of small RNA abundance was calculated by 21-nt or 24-nt reads number falling into 1 kb windows/regions throughout the whole genomes and windows containing more than 3 reads were tested by EdgeR (Robinson et al., 2010) between samples. Windows with corrected p-value ≤0.01, were defined as differential smRNA regions.
ChIP-seq's peaks calling were used with MACS software with default parameters.
The Differentially Methylated Cytosines (DMCs) between any two genotypes were defined by binomial test (methylSig) (Park et al., 2014). Only cytosine sites whose adjusted q-values ≤ 0.01 and DNA methylation difference of 0.7, 0.5 and 0.1 between any two genotypes were designated for DMCs (Stroud et al., 2013). The Differentially Methylated Regions (DMRs), Differentially Methylated Genes (DMGs) were calculated as DMCs.
Genome_build: MSU7.0: bulid_all genome.fa
Supplementary_files_format_and_content: wig and peaks' xls files were generated using MACS software;
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| Submission date |
May 13, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
chao zhou |
| E-mail(s) |
zhouchao@ctgu.edu.cn
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| Organization name |
CTGU
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| Street address |
No.8,daxuelu Street
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| City |
yichang |
| ZIP/Postal code |
443002 |
| Country |
China |
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| Platform ID |
GPL13834 |
| Series (1) |
| GSE81436 |
The ortholog of DDM1 is mainly required for CHG and CG methylation of heterochromatin and is involved in DRM2-mediated CHH methylation that targets mostly genic regions of the rice genome |
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| Relations |
| BioSample |
SAMN04999533 |
| SRA |
SRX1760739 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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