|
Status |
Public on Mar 18, 2014 |
Title |
Col-0 (bisulfite-seq) |
Sample type |
SRA |
|
|
Source name |
Arabidopsis stages 9 to 12 inflorescence
|
Organism |
Arabidopsis thaliana |
Characteristics |
tissue: plant inflorescence genotype/variation: Col-0
|
Growth protocol |
Arabidopsis grown on soil under standard long day growth conditions.
|
Extracted molecule |
genomic DNA |
Extraction protocol |
DNA was isolated from pooled inflorescence tissue (n = 3) collected phenol/chloroform extraction. For each library, 1-5 μg of genomic DNA was sheared using a Covaris S220 Adaptive Focused Acoustics ultra sonicator. Libraries were constructed following standard protocol using the NEB Next DNA Sample Prep Master Mix Set 1 (NEB E6040) and Illumina-compatible paired-end adaptors, which had all cytosines, methylated. 50 ng of each library was treated with sodium bisulphite using EZ DNA Methylation-GoldTM Kit (Zymo Research D5005) according to the protocol provided by the manufacturer. For each purified library 5-10 ng of purified library was amplified using Expand High Fidelity PLUS PCR system (Roche 03300242001), which is capable of efficiently amplifying uracil-containing templates. 50 μl reactions contained 200 μM each dNTP, 1 μM primer, 2.5 mM MgCl2 and 2.5 U Expand HiFi enzyme, and were performed according to the manufacturer’s instructions for 18 cycles. Amplified libraries were ran on 2% MetaPhor® agarose (Lonza 50108) gel. Fragments of 220-350 bp were excised from gel and purified using the QIAquick PCR Purification kit (Qiagen 28104). DNA concentrations were quantified on Bioanalyzer (Agilent), diluted to 10 nM and loaded on flow cells to generate clusters.
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|
|
Library strategy |
Bisulfite-Seq |
Library source |
genomic |
Library selection |
RANDOM |
Instrument model |
Illumina Genome Analyzer |
|
|
Description |
DNA methylation analysis in Arabidopsis
|
Data processing |
Base call and quality scores were generated using the standard Illumina Pipeline for Paired-End Reads. BS-seq reads were trimmed for adaptor sequence and low quality base call, then aligned to ZmB73 4a.53 genome assembly using rmap V2.03 with parameters -Q -v -m 4 Then the C/T counts from aligned BS-seq reads for each cytosine in reference genome were saved in CT count file Genome_build: TAIR10 Supplementary_files_format_and_content: The CT count files have the first 6 columns the same as bed file, the 7th col is count of C, the 8th col is count of T, the 9th col is the 3 nt in reference genome, the 10th col is the context of cytosine
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|
|
Submission date |
Nov 13, 2013 |
Last update date |
May 15, 2019 |
Contact name |
Kate M Creasey |
Organization name |
Cold Spring Harbor Laboratory
|
Department |
Plant Sciences
|
Street address |
1 Bungtown Road
|
City |
Cold Spring Harbor |
State/province |
NY |
ZIP/Postal code |
11724 |
Country |
USA |
|
|
Platform ID |
GPL9062 |
Series (2) |
GSE52346 |
miRNAs trigger widespread epigenetically-activated siRNAs from transposons in Arabidopsis (bisulfite-seq) |
GSE52952 |
miRNAs trigger widespread epigenetically-activated siRNAs from transposons in Arabidopsis |
|
Relations |
BioSample |
SAMN02422663 |
SRA |
SRX384340 |