library_construction_protocol: The sperm collected by FACS was lysed and then RNA was enriched for sequencing. RNAs were diluted to 0.2 ng/μL and 2.5 μL samples were performed library constructions. The samples were incubated with oligo(dT) primers (a deoxynucleoside triphosphate mixture) for 3 min at 72 °C, and cDNA pools were obtained by Smart-seq2 reverse transcription reactions. After the first-strand reaction, the cDNAs were pre-amplified with limited cycles (~ 12 cycles). Sequencing libraries were then constructed with 0.5 ng cDNA using the TruePrep DNA Library Prep Kit V2 for Illumina (TD503; Vazyme, Nanjing, China) according to the manufacturer's instructions. Barcoded libraries were pooled and sequenced on the Illumina NovaSeq 6000 platform in the 151 bp paired-end mode.