|
| Status |
Public on Aug 17, 2021 |
| Title |
C-OE2-2 |
| Sample type |
SRA |
| |
|
| Source name |
Tobacco leaf blades
|
| Organism |
Nicotiana tabacum |
| Characteristics |
tissue: The 5th leaf blades (with main veins removed)
developmental stage: About 34 days after germination
genotype: NtCBL5A-OE
treatment: Control
|
| Treatment protocol |
Tobacco was treated under 100 mM NaCl for 4 days
|
| Growth protocol |
Salt stress experiments were conducted in 2019 at Unifarm, Wageningen University & Research in the Netherlands. Conditions of the greenhouse were 16 h light / 8 h dark at 25/23℃ and 70% relative humidity. The shortwave radiation level was maintained in the greenhouse compartment using artificial PAR (photosynthetically active radiation) when the incoming shortwave radiation was below 200 Wm-2. For the salt tolerance evaluations, tobacco seeds were sown in soil, and they germinated after about 8 days. About 12 days after germination (DAG), the young seedlings were transplanted in rock-wool plugs within float trays for 8 days, then they were transplanted to a circular flow hydroponic system filled with 1/2 Hoagland’s nutrient solution (500 L). The water used to prepare 1/2 Hoagland’s nutrient solution contained trace amounts of Na+ and Cl- (5.04 μg/ml and 6.72 μg/ml, respectively). After 6 days of acclimatization (at ~ 30 DAG), NaCl was added to the nutrient solution to a concentration of 50 mM on the first day to avoid salt shock, and the final NaCl concentration of 100 mM NaCl was reached the next day.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
At 4 DAT, the 5th leaf blades (with main veins removed) of four individual plants under control conditions and saline conditions in the daytime were sampled and frozen immediately in liquid nitrogen. Leaf blades from four individual plants were mixed and ground to powder. RNA was isolated and purified with RNeasy Plus Mini Kit (Qiagen, Cat No./ID: 74134, the Netherlands) following the manufacturer’s protocol. RNA samples of WT and OE-2 overexpression lines from two independent salt treatment experiments were used for transcriptome sequencing.
RNA libraries were prepared for sequencing using standard Illumina protocols
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
RNA-seq was performed by the Novogene using Illumina Polymerase-based sequencing-by-synthesis, obtaining a read length of 150 bp and coverage of 48 to 81 million reads per sample.
Raw reads of fastq format were firstly processed through in-house perl scripts so that all the downstream analyses were based on the clean data with high quality.
Reference genome and gene model annotation files (ftp://ftp.solgenomics.net/genomes/Nicotiana_tabacum/edwards_et_al_2017/assembly/Nitab-v4.5_genome_Chr_Edwards2017.fasta.gz ) were downloaded from NCBI.
Index of the reference genome was built using Hisat2 v2.0.5 and paired-end clean reads were aligned to the reference genome using Hisat2 v2.05.
The mapped reads of each sample were assembled by StringTie (v1.3.3b) (Pertea et al., 2015) in a reference-based approach.
FeatureCounts v1.5.0-p3 was used to count the reads numbers mapped to each gene and then FPKM of each gene was calculated based on the length of the gene and read counts mapped to this gene.
DEGs of two groups were performed using the DESeq2 R package (1.20.0) and resulting P-values were adjusted using the Benjamini and Hochberg’s approach for controlling the false discovery rate.
Genes with an adjusted P-value <0.05 found by DESeq2 were assigned as differentially expressed. ClusterProfiler R package was used to test the statistical enrichment of DEGs in KEGG pathways.
Genome_build: Nitab4.5
Supplementary_files_format_and_content: tab-delimited text files include readcount and FPKM values for each Sample ...
|
| |
|
| Submission date |
Jul 30, 2021 |
| Last update date |
Aug 17, 2021 |
| Contact name |
Qian Wang |
| E-mail(s) |
wangqian01@caas.cn
|
| Organization name |
Chinese Academy of Agricultural Sciences
|
| Street address |
Keyuanjingsi Road, Laoshan District
|
| City |
Qingdao |
| State/province |
Shandong |
| ZIP/Postal code |
266101 |
| Country |
China |
| |
|
| Platform ID |
GPL29390 |
| Series (1) |
| GSE181164 |
Differentially Expressed Genes in Wild-type Tobacco Leaves and NtCBL5A-OE Transgenic Tobacco Leaves Under Salt Stress |
|
| Relations |
| BioSample |
SAMN20501033 |
| SRA |
SRX11609800 |
