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Sample GSM4367757

Query DataSets for GSM4367757

Status

Public on Mar 01, 2022

Title

NT24WT2

Sample type

SRA

Source name

normal temperature from ZT20-ZT24_seedlings

Organism

Oryza sativa

Characteristics

cultivar: 93-11 (O. sativa ssp. indica)
developmental stage: three-leaf stage
tissue: above ground part of seedlings
treatment: 4 hrs treatment of normal temperature (28°C) from ZT20-ZT24 before harvesting

Treatment protocol

10-day seedlings at three-leaf stage were treated with chilling temperature (13°C) for 4 hrs at different Zeitgeber Time (ZT) (ZT0-ZT36) before harvesting for RNA extraction on the 11th day. For transcriptome analyses, CT12 group seedlings experienced 4 hrs treatment of chilling temperature from ZT8-ZT12, while NT12 group seedlings were kept at 28°C before harvesting at ZT12 as controls. CT24 group seedlings were chilling-treated from ZT20 to ZT24 before being harvested, while NT24 group seedlings were harvested at the same time at normal growth temperature. The samples were immediately frozen in liquid nitrogen and stored in –80°C freezer until RNA extraction.

Growth protocol

Rice cultivar “93-11” (O. sativa ssp. indica) was used for this study. Seeds were soaked in water for 36 hrs and then incubated at 30 °C for 6 days. The germinated seeds were sown in a round pot (diameter 8cm) for RNA sampling (25 seedlings per pot). The seedlings were incubated in 12 h light/12 h dark cycles (12L:12D) at 28 °C (white light intensity: 248 μmol m–2 s–1; relative humidity: 70 ± 5%) for about 10 days until the three-leaf stage, in a growth chamber (R4630, Ningbo Dongnan Instrument Co., Ltd., China).

Extracted molecule

total RNA

Extraction protocol

Total RNA was extracted from up-ground rice seedlings using Trizol (Invitrogen, Carlsbad, CA, USA) following manufacturer's guidelines. For each treatment, RNA extraction was performed in three biological repeats (5 plants per repeat). NanoPhotometer® (IMPLEN, CA, USA) and Qubit® RNA Assay Kit in a Qubit® 2.0 Fluorometer (Life Technologies, CA, USA) were used for purity and quantitative assessment of RNA samples, respectively. In addition, the integrity of RNA was assessed using the RNA Nano 6000 Assay Kit of the Bioanalyzer 2100 system (Agilent Technologies, CA, USA).
Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (NEB, USA) according the manufacturer’s instructions and index codes were added to attribute sequences to each sample.

Library strategy

RNA-Seq

Library source

transcriptomic

Library selection

cDNA

Instrument model

Illumina HiSeq 4000

Data processing

Illumina Casava1.8 software used for basecalling.
Raw data (raw reads) of fastq format were firstly processed through in-house perl scripts.In this step, clean data (clean reads) were obtained by removing reads containing adapter, reads containing ploy-N and low quality reads from raw data. At the same time, Q20, Q30 and GC content the clean data were calculated. All the downstream analyses were based on the clean data with high quality.
Filtered reads were aligned to the reference rice genome (Oryza sativa R498_IGDBv3_coreset_MSU) using HISAT(2.0.4).
Genome_build: Oryza sativa R498_IGDBv3_coreset_MSU
Supplementary_files_format_and_content: a text file include count number for each sample.

Submission date

Mar 02, 2020

Last update date

Mar 01, 2022

Contact name

Xuedan Lu

E-mail(s)

luxuedan1@126.com

Phone

+8615388075635

Organization name

Hunan Agricultural University