|
| Status |
Public on Feb 10, 2022 |
| Title |
AT12_scRNA-seq |
| Sample type |
SRA |
| |
|
| Source name |
Arabidopsis thaliana seedings
|
| Organism |
Arabidopsis thaliana |
| Characteristics |
tissue: the wounded region of leaf explants
genotype: wild type col-0
Stage: 4 d after detachment
|
| Treatment protocol |
The first pair of rosette leaves were cut at the junction of the blade and petiole, and the detached blades were cultured on sucrose-free B5 medium at 22°C under 24-h light conditions for 4 d .
|
| Growth protocol |
Arabidopsis Col-0 were germinated and grown on 1/2 MS medium with 1% sucrose at 22°C under a 16-h light/8-h dark photoperiod for 12 d.
|
| Extracted molecule |
polyA RNA |
| Extraction protocol |
The base region of ~50 leaves at 4 d after detachment were harvested and digested for 2 h at room temperature in RNase-free enzyme solution (1.5% cellulase R10, 1.2% macerozyme R10, 0.4 M mannitol, 3 mM -mercaptoethanol, 10 mM CaCl2 and 0.3% BSA). The protoplast solution was strained through a 40-mm filter and a 30-mm filter. The filtered solution was centrifuged at 500 g for 3 min, and washed twice with 8% mannitol at room temperature. The protoplast pellet was resuspended in 50 μl of 8% mannitol with 0.04% BSA. About 94% cell viability was confirmed by trypan blue staining. Then the single cell suspension was counted using the hemocytometer and the concentration was adjusted to 500 cells/μl.
Single-cell suspension was loaded to 10× Chromium to capture 8,000 single cells according to the manufacturer’s instructions of 10× Genomics Chromium Single-Cell 3’ kit (V3, 10× Genomics, USA) .The library was constructed according to the standard protocol and deep sequencing was performed by Illumina NovaSeq 6000 sequencing (paired-end multiplexing run,150 bp) (LC-Bio Technology, China).
|
| |
|
| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
| |
|
| Data processing |
For single cell RNA-seq, raw data was mapped to Arabidopsis thaliana genome (TAIR10) using Cell Ranger v3.1.0 with parameter “expect-cells=5,000”. gene expression matrix from Cell Ranger was input to R package Seurat v3.0.2 which was used for cell clustering and dimensionality reduction analysis.
Supplementary_files_format_and_content: tab-delimited text files include UMI for each gene in each cell
|
| |
|
| Submission date |
Mar 20, 2020 |
| Last update date |
Feb 10, 2022 |
| Contact name |
Lin XU |
| E-mail(s) |
xulin01@sibs.ac.cn
|
| Phone |
86-21-54924101
|
| Organization name |
Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences
|
| Department |
Chinese Academy of Sciences
|
| Street address |
300 Fenglin Road
|
| City |
Shanghai |
| ZIP/Postal code |
200032 |
| Country |
China |
| |
|
| Platform ID |
GPL26208 |
| Series (1) |
| GSE147289 |
Single-Cell RNA-seq analysis of cell lineages in De Novo Root Regeneration |
|
| Relations |
| BioSample |
SAMN14415354 |
| SRA |
SRX7960932 |
