GSM7068207
Lateral root tips of 0.5–1-cm were cut into pieces with a sharp blade and placed in a six-well cell culture dish containing 10 mL zymolytic solution (1.5% cellulase R10, 1% pectolyase, 0.4 M mannitol, 0.1 M KCl, 0.08 M MES, 0.02 M CaCl2 and 0.1% BSA). Subsequently, vacuum treatment (0.05 MPa) was conducted under a dark environment for 1 h, and the enzymatic hydrolysis reaction lasted for about 6 h in a constant-temperature shaker with a gentle speed of 80 r/min at 25°C. A 40-μm cell strainer was used to filter the dissociated protoplasts. The protoplasts were collected by centrifugation and washed 2–4 times with WB buffer (0.1 M KCl, 0.08 M MES, 0.4 M mannitol and 0.1% BSA) on standby. The 10× Genomics 3ʹ Single Cell Platform was adopted in this study to process the high-quality protoplasts according to the corresponding instructions, and single-cell gel beads in emulsions (GEMs) were generated by a microfluidic chip in different lanes, and the scRNA-seq library was then constructed using Chromium Next GEM Single Cell 3ʹ Reagent Kits v3.1 (10× Genomics, Pleasanton, CA).