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| Status |
Public on Jun 26, 2017 |
| Title |
Nb A&Z |
| Sample type |
SRA |
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| Source name |
Aerial part of 10-day-old seedlings, alpha-amanitin and zebularine
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| Organism |
Oryza sativa Japonica Group |
| Characteristics |
cultivar: Nipponbare
tissue: Aerial part of seedlings
age: 10 days
treatment: alpha-amanitin and zebularine
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| Treatment protocol |
Treatment with alpha-amanitin and zebularine: Germination and growth on solid ½ MS medium (1% sucrose, 0.5% Phytagel (Sigma), pH 5.8) supplemented with sterile filtrated zebularine (Sigma, stock: 5 mg/ml in DMSO) and α-amanitin (5 µg/ml) (Sigma, stock: 1 mg/ml in water).
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| Growth protocol |
Germination and growth for 10 days at 12 h at 28 °C (day) and 27 °C (night) on solid ½ MS medium (1% sucrose, 0.5% Phytagel (Sigma), pH 5.8).
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Total DNA of seedlings was extracted as previously reported (Mette, M. F. et al. (1999) EMBO J 18, 241-248). Enrichment of extrachromosomal circular DNA was performed according to (Lanciano et al. submitted). 5 μg of genomic DNA for each sample were purified using a Geneclean kit (MPBio, USA) according to the manufacturer's instructions. ecDNA was isolated from the GeneClean product using the PlasmidSafe DNase (Epicentre, USA) according to the manufacturer's instructions, except that the 37°C incubation was performed for 17h. DNA samples were precipitated by adding 0.1 volume of 3M sodium acetate (pH 5.2), 2.5 volumes of ethanol and 1 μl of glycogen (Fisher, USA) and incubating overnight at -20°C. The precipitated circular DNA was amplified by random rolling circle amplification using the Illustra TempliPhi kit (GE Healthcare, USA) according to the manufacturer's instructions except that the incubation was performed for 65h at 28°C. The DNA concentration was determined using the DNA PicoGreen kit (Invitrogen, USA) using a LightCycler480 (Roche, USA).
One nanogram of amplified ecDNA from each sample was used to prepare the libraries using the Nextera XT library kit (Illumina, USA) according to the manufacturer's instructions.
Samples were pooled and loaded onto a MiSeq platform (Illumina, USA) and 2x250 nucleotides paired-end sequencing was performed.
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| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina MiSeq |
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| Description |
Sample name: NB19
Oryza sativa japonica cv. Nipponbare
Extrachromosomal circular-DNA enriched genomic DNA
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| Data processing |
Quality control of FASTQ files was evaluated using the FastQC tool (version 0.10.1).
To remove any read originating from organelle circular genomes, reads were mapped against the mitochondria and chloroplast genomes using the program Bowtie2 version 2.2.2 71 with --sensitive local mapping.
Unmapped reads were mapped against the reference genome IRGSP-1.0 (http://rgp.dna.affrc.go.jp/E/IRGSP/Build5/build5.html) using the following parameters: --sensitive local, -k 1. DNA from both mitochondria and chloroplast genomes integrated in nuclear genomes was masked (1,697,400 bp). The TE containing regions cover 194,224,800 bp in O. sativa.
Genome_build: IRGSP-1.0
Supplementary_files_format_and_content: bigWig files
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| Submission date |
Nov 23, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Etienne Bucher |
| E-mail(s) |
etienne.bucher@angers.inra.fr
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| Phone |
++33 2 41 22 57 97
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| Organization name |
IRHS
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| Lab |
Bucher
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| Street address |
42 rue Georges Morel
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| City |
Beaucouzé |
| ZIP/Postal code |
49071 |
| Country |
France |
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| Platform ID |
GPL22703 |
| Series (1) |
| GSE90484 |
Inhibition of RNA polymerase II allows controlled mobilisation of retrotransposons for plant breeding [rice] |
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| Relations |
| BioSample |
SAMN06052015 |
| SRA |
SRX2372569 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2402734_NB19.bigwig |
2.3 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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