*Article* **Transcriptome Analysis Reveals the Genes Related to Pollen Abortion in a Cytoplasmic Male-Sterile Soybean (***Glycine max* **(L.) Merr.)**

**Zhiyuan Bai 1,2,†, Xianlong Ding 1,†, Ruijun Zhang <sup>2</sup> , Yuhua Yang <sup>2</sup> , Baoguo Wei <sup>2</sup> , Shouping Yang 1,\* and Junyi Gai 1,\***


**Abstract:** Cytoplasmic male sterility (CMS) lays a foundation for the utilization of heterosis in soybean. The soybean CMS line SXCMS5A is an excellent CMS line exhibiting 100% male sterility. Cytological analysis revealed that in SXCMS5A compared to its maintainer SXCMS5B, its tapetum was vacuolated and abnormally developed. To identify the genes and metabolic pathways involving in pollen abortion of SXCMS5A, a comparative transcriptome analysis was conducted between SXCMS5A and SXCMS5B using flower buds. A total of 372,973,796 high quality clean reads were obtained from 6 samples (3 replicates for each material), and 840 differentially expressed genes (DEGs) were identified, including 658 downregulated and 182 upregulated ones in SXCMS5A compared to SXCMS5B. Among them, 13 DEGs, i.e., 12 open reading frames (ORFs) and 1 *COX2*, were mitochondrial genome genes in which *ORF178* and *ORF103c* were upregulated in CMS lines and had transmembrane domain(s), therefore, identified as CMS candidate mitochondrial genes of SXCMS5A. Furthermore, numerous DEGs were associated with pollen wall development, carbohydrate metabolism, sugar transport, reactive oxygen species (ROS) metabolism and transcription factor. Some of them were further confirmed by quantitative real time PCR analysis between CMS lines with the same cytoplasmic source as SXCMS5A and their respective maintainer lines. The amount of soluble sugar and adenosine triphosphate and the activity of catalase and ascorbic acid oxidase showed that energy supply and ROS scavenging decreased in SXCMS5A compared to SXCMS5B. These findings provide valuable information for further understanding the molecular mechanism regulating the pollen abortion of soybean CMS.

**Keywords:** soybean (*Glycine max* (L.) Merr.); cytoplasmic male sterility; pollen abortion; gene expression; RNA sequencing
