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Sample GSM8209525

Query DataSets for GSM8209525

Status

Public on Apr 17, 2024

Title

WT_WW_14

Sample type

SRA

Source name

Leaf

Organism

Solanum lycopersicum

Characteristics

cultivar: M82
tissue: Leaf
genotype: WT
treatment: Well-watered
time: T0

Treatment protocol

Time-course experiments were performed by gently uprooting plants of both wt and SL- genotypes, transferring them in vermiculite hydrated with only 20 ml of water (stressed group), or transferred in wet vermiculite (well-watered controls). After having completely recovered, the stressed plants were subjected to a second identical cycle of dehydration and rewatering. Unstressed, well-watered controls were sampled before transfer to new wet substrates. GR245DS treatment was carried out by leaf spraying of a 5 uM solution until drip-off 24 h before sampling, which corresponded with the beginning of the drought time-course (hence, untreated irrigated wt plants are an appropriate control group for this condition as well).

Growth protocol

Seeds were sterilized in 4% (v/v) sodium hypochlorite containing 0.02% (v/v) Tween 20, rinsed thoroughly with sterile water, and then placed for 48 h on moistened filter paper at 25°C in darkness. Germinated seedlings were grown for two weeks in a walk-in climate chamber (16/8h light/dark 25°C, 65% humidity, and 200 µmol s–1 m–2 of photosynthetic photon flux density) in a seedbed with standard soil (Terra Nature, NPK 12:14:24) and then moved into 1-liter pots filled with vermiculite and kept under the same conditions.

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted from leaves using the Spectrum Plant Total RNA Kit (Sigma Aldrich). After digestion of contaminant DNA by DNAse I (ThermoScientific) at 37°C for 30 min, RNA quantity and quality were determined with a Nanodrop 2000 spectrophotometer (Thermo Fisher Scientific Inc., Wilmington, DE, United States).
cDNA libraries were prepared from 1 µg total RNA using NEBnext Ultra TM RNA Library Prep Kit for Illumina (NEB, USA) following the manufacturer’s recommendations. A total of 21 libraries (three each for wt and SL- leaves, plus wt treated with GR245DS) were constructed and quantified using a Qubit 2.9 fluorometer (Life Technologies) and sequenced on an Illumina platform to generate paired-end reads.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina NovaSeq 6000

Data processing

Clean reads were obtained by removing reads containing adapter and poly-N sequences and reads with low quality from raw data. At the same time, Q20, Q30 and GC content of the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files were downloaded from genome website browser (NCBI/UCSC/Ensembl) directly. Paired-end clean reads were mapped to the reference genome (SL3.0) using the HISAT2 software.
HTSeq was used to count the read numbers mapped of each gene, including known and novel genes; then RPKM of each gene was calculated based on the length of the gene and reads count mapped to this gene.
Assembly: ITAG SL3.0
Supplementary files format and content: tab-delimited text file includes raw counts for each sample

Submission date

Apr 16, 2024

Last update date

Apr 17, 2024

Contact name

IVAN VISENTIN

E-mail(s)

plantstresslab2024@gmail.com

Phone

+390116708805

Organization name

University of Turin