repeat2_long
Arabidopsis thaliana
Protocols: For isolation of leaf protoplasts (~5 excised leaves from four-week-old seedlings pooled per sample), a “tape-sandwich” method was used, where the abaxial epidermis of leaves was peeled using adhesive tape (Scotch® Magic™, 3M) and leaves were immediately immersed in a cell-wall degrading enzymatic buffer in protoplasting buffer (0.6 M mannitol, 20 mM KCl, 10 mM CaCl2, 20 mM MES, 0.1% BSA, 1.0% cellulase R10, 0.3% macerozyme) at pH 5.7. The enzymatic reaction was performed for 1 h at room temperature with gentle rotation (30 rpm) in the dark. The solution was filtered through pre-wet 70 μm nylon mesh, collected by centrifugation at 200 × g for 6 min and resuspended in wash buffer (0.6 M mannitol, 20 mM KCl, 10 mM CaCl2, 20 mM MES). Arabidopsis (Arabidopsis thaliana L. Heynh.) cv. Columbia-0 (Col-0) was grown in a controlled-environment growth chamber (Weiss Technik).  Beads in emulsion (GEM) generation and droplet barcoding were performed according to the 10x Genomics protocols with Chromium Next GEM Single Cell 3'Kit v3.1 and Chromium Next GEM Chip G Single Cell Kit, aiming for 10,000 cells per sample. Unfragmentated cDNA was prepared for nanopore sequencing on the ONT PromethION (P24) with a combination of the primers from SQK-PCS111, rapid adapters from EXP-RAA114, and auxiliary vials for loading on R10.4.1 flow cells from EXP-AUX003.
ENA first public
2026-02-27
INSDC center name
VIB Single Cell Core
INSDC status
public
Submitter Id
E-MTAB-13866:repeat2_long
age
4
broker name
ArrayExpress
cell_type
all
collection date
not collected
common name
thale cress
developmental stage
mature leaf
genotype
wild type genotype
geographic location (country and/or sea)
not collected
isolate
not applicable
organism part
leaf
replicate
2
sample name
E-MTAB-13866:repeat2_long
scientific_name
Arabidopsis thaliana
ecotype
Col-0