protoplasts of root tips
The Chromium Next GEM Single cell 3 Reagent v3.1 Kit was used for the single cell RNA-seq. A total of 16,000 sorted cells were loaded on the 10x Genomics microfluidic chip with RT-PCR master mix, barcoded Single-Cell 3 v3.1 Gel Beads and Partitioning Oil to generate single cell beads in emulsions (GEMs). Reverse-transcription was performed in droplets after gel beads dissolving to produce barcoded full-length cDNAs. After incubation, GEMs were disrupted and cDNAs were cleaned up with DynaBeads Myone Silane Beads before being amplified by 12 cycles of PCR. Finally, amplified cDNAs were fragmented, end-repaired, A-tailed and index adaptor-ligated. The final libraries were amplified and sequenced on the Illumina sequencing platform.