Single-cell RNA sequencing of Schizosaccharomyces Pombe: Ancestor, h90, 10h under nitrogen starvation
Schizosaccharomyces pombe strains (Y447, prm1 Ancestor) were cultured in MSL-N medium and harvested at designated time points (10h) during mating in liquid culture or on agar plates under nitrogen starvation. For single-cell RNA-seq (scRNA-seq), cell walls were removed using 0.2 g/mL Lallyzyme following sonication (60W) to generate protoplasts, which were then quantified and loaded onto SCOPE-chip HD microfluidic chips. Single-cell transcriptomic libraries were constructed using the GEXSCOPE Microbial Single Cell RNA Library Kit HD (Yeast) (Singleron Biotech) according to the manufacturers protocol, involving mRNA capture via Barcode Beads HD V3 by hybridizing to their poly(A) tails, reverse transcription, and cDNA amplification. The final libraries were sequenced on the DNBSEQ-T7 platform (MGI) to generate paired-end reads. Parallel bulk RNA-seq samples were harvested and snap-frozen in liquid nitrogen for storage at -80C.