Single-cell RNA sequencing (scRNA-seq) is a high-tech method for characterizing the expression patterns of heterogeneous cells in the same tissue and has changed our evaluation of biological systems by increasing the number of individual cells analyzed. However, the full potential of scRNA-seq, particularly in plant science, has not yet been elucidated. To explore the utilization of scRNA-seq technology in plants, we firstly conducted a comprehensive review of significant scRNA-seq findings in the past few years. Secondly, we introduced the research and applications of scRNA-seq technology to plant tissues in recent years, primarily focusing on model plants, crops, and wood. We then offered five databases that could facilitate the identification of distinct expression marker genes for various cell types. Finally, we analyzed the potential problems, challenges, and directions for applying scRNA-seq in plants, with the aim of providing a theoretical foundation for the better use of this technique in future plant research.

The cytoplasmic male sterility/restorer-of-fertility (CMS/Rf) system plays a vital role in high-efficiency hybrid seed production in crops, including soybean (Glycine max (L.) Merr.). The markers linked to fertility restoration and the restorer-of-fertility (Rf) genes are essential because they can facilitate the breeding of new CMS lines and production of commercial hybrid soybean seeds. To date, several soybean Rf genes have been mapped to various genetic loci in diverse genetic populations. However, the mapping range of restorer genes remains narrow, with relatively limited practical applicability. Therefore, in the present study, F₂ and F₃ segregating populations derived from the CMS line JLCMS5A crossed with the restorer line JLR2 were developed and used for Rf3 gene fine mapping. Genetic investigation indicated that the restorer line JLR2 was controlled by a single dominant gene, Rf3. By integrating bulk-segregant analysis and next-generation sequencing, a 4 Mb region on chromosome 9 was identified, which was most likely the target region harboring the candidate gene responsible for fertility restoration. This region was further narrowed down to 86.44 Kb via fine mapping in F₂ and F₃ populations using SSR, InDel, and dCAPS markers. This region contained 10 putative genes (Glyma.09G171100–Glyma.09G172000). Finally, Glyma.09G171200, which encodes a mitochondria-targeted pentatricopeptide repeat protein, was proposed as the potential candidate for Rf3 using sequence alignment and expression analysis in restorer and CMS lines. Based on single-nucleotide polymorphisms in Glyma.09G171200, a CAPS marker co-segregated with Rf3 named CAPS1712 was developed. Our results will be fundamental in the assisted selection and creation of potent lines for the production and rapid selection of novel restorer lines.

(1) Background: Cytoplasmic male sterility (CMS) is important for exploiting heterosis. Soybean (Glycine max L.) has a low outcrossing rate that is detrimental for breeding sterile lines and producing hybrid seeds. Therefore, the molecular mechanism controlling the outcrossing rate should be elucidated to increase the outcrossing rate of soybean CMS lines; (2) Methods: The male–sterile soybean lines JLCMS313A (with a high outcrossing rate; HL) and JLCMS226A (with a low outcrossing rate; LL) were used for a combined analysis of the transcriptome (RNA–seq) and the targeted phenol metabolome; (3) Results: The comparison between HL and LL detected 5946 differentially expressed genes (DEGs) and 81 phenolic metabolites. The analysis of the DEGs and differentially abundant phenolic metabolites identified only one common KEGG pathway related to flavonoid biosynthesis. The qRT–PCR expression for eight DEGs was almost consistent with the transcriptome data. The comparison of the cloned coding sequence (CDS) regions of the SUS, FLS, UGT, and F3H genes between HL and LL revealed seven single nucleotide polymorphisms (SNPs) only in the F3H CDS. Moreover, five significant differentially abundant phenolic metabolites between HL and LL were associated with flavonoid metabolic pathways. Finally, on the basis of the SNPs in the F3H CDS, one derived cleaved amplified polymorphic sequence (dCAPS) marker was developed to distinguish between HL and LL soybean lines; (4) Conclusions: The flavonoid biosynthesis pathway may indirectly affect the outcrossing rate of CMS sterile lines in soybean.