|
| Status |
Public on Sep 06, 2016 |
| Title |
Ws ecotype Arabidopsis seed 17daf |
| Sample type |
SRA |
| |
|
| Source name |
seed
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
genotype/variation: Wildtype
ecotype: Wassilewskija
developmental stage: 17daf
phenotype: Desiccation tolerant seed
|
| Growth protocol |
Plants of A. thaliana were grown in a sterile mix of vermiculite and soil in a growth chamber at 22 °C with a 16-h photoperiod at 200 mmol m2 s. the flowers were marked, and at given time intervals following anthesis (15, 17 and 21 DAF), siliques were collected from 24 plants, seeds were dissected, immediately frozen in liquid nitrogen, and stored at -80°C. Samples were collected in triplicates
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For RNA-Seq total RNA was isolated using the TRIZOL reagent (Invitrogen) according to the manufacturer's instructions and further purificated using the Qiagen RNeasy plant mini kit according to the manufacturer's instructions.
Five μg of RNA from each experimental condition was pooled to obtain a single RNA sample. First and second strand cDNA synthesis was performed with 3 μg of the total RNA mixture using Message Amp-II kit (Ambion) following the manufacturers recommendations. Each sample was treated separately 10-12 ng of synthesized cDNA was amplified by in vitro transcription and the resulting 70-90 μg of antisense RNA (aRNA) purified using RNAeasy columns (Qiagen). A second round of cDNA synthesis was performed using the mRNA as template (20 μg). cDNA synthesis was performed as described above except that random primers (mostly hexamers) were used at the first strand synthesis stage. This procedure yielded approximately 10 μg of cDNA that was purified using the DNA Clear Kit for cDNA purification (Ambion). Samples were barcoded for multiplexing using the SOLiD RNA Barcoding Kit
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
AB SOLiD 4 System |
| |
|
| Description |
ws_17daf
|
| Data processing |
The SOLiD BioScope Whole Transcriptome Analysis (WTA) version 1.2.1 pipeline for single reads was used to align the reads to the Col-0 reference genome TAIR10
Two perfect matches per location were allowed, count aligned reads per exon, and calculate per base coverage. WTA pipeline CountTags module provides normalize RPKM (reads per kilobase of exon sequence, per million reads) values along with read counts per exon. Using BASH and MySQL scrips were calculated the gene counts per gene model
Gene counts were normalized using edgeR’s TMM [trimmed mean M (¼ log fold-change gene expression)] algorithm
Genome_build: TAIR 10
Supplementary_files_format_and_content: counts_libraries_matrix.csv: raw counts of sequencing reads
Supplementary_files_format_and_content: cpm_normalized_edgeR.csv: normalized counts of sequencing reads
|
| |
|
| Submission date |
Dec 15, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Sandra Isabel Gonzalez-Morales |
| E-mail(s) |
sgonzalez@langebio.cinvestav.mx
|
| Organization name |
CINVESTAV
|
| Department |
LANGEBIO
|
| Street address |
Km 9.6 Libramiento Norte Carretera León
|
| City |
Irapuato |
| State/province |
Guanajuato |
| ZIP/Postal code |
36821 |
| Country |
Mexico |
| |
|
| Platform ID |
GPL14599 |
| Series (1) |
| GSE76015 |
Desiccation Tolerance of Arabidopsis thaliana seeds |
|
| Relations |
| BioSample |
SAMN04338094 |
| SRA |
SRX1482502 |
