Plant mitochondrial (mt) genomes are species specific due to the vast of foreign DNA migration and frequent recombination of repeated sequences. Sequencing of the mt genome of kenaf (Hibiscus cannabinus) is essential for elucidating its evolutionary characteristics. In the present study, single-molecule real-time sequencing technology (SMRT) was used to sequence the complete mt genome of kenaf. Results showed that the complete kenaf mt genome was 569,915 bp long and consisted of 62 genes, including 36 protein-coding, 3 rRNA and 23 tRNA genes. Twenty-five introns were found among nine of the 36 protein-coding genes, and five introns were trans-spliced. A comparative analysis with other plant mt genomes showed that four syntenic gene clusters were conserved in all plant mtDNAs. Fifteen chloroplast-derived fragments were strongly associated with mt genes, including the intact sequences of the chloroplast genes psaA, ndhB and rps7. According to the plant mt genome evolution analysis, some ribosomal protein genes and succinate dehydrogenase genes were frequently lost during the evolution of angiosperms. Our data suggest that the kenaf mt genome retained evolutionarily conserved characteristics. Overall, the complete sequencing of the kenaf mt genome provides additional information and enhances our better understanding of mt genomic evolution across angiosperms.

Cytoplasmic male sterility (CMS) is a maternally inherited trait used for hybrid production in plants, a novel kenaf CMS line 722HA was derived from the thermo-sensitive male-sterile mutant ‘HMS’ by recurrent backcrossing with 722HB. The line 722HA has great potential for hybrid breeding in kenaf. However, the underlying molecular mechanism that controls pollen abortion in 722HA remains unclear, thus limiting the full utilization of this line. To understand the possible mechanism governing pollen abortion in 722HA, cytological, transcriptomic, and biochemical analyses were carried out to compare the CMS line 722HA and its maintainer line 722HB. Cytological observations of the microspore development revealed premature degradation of the tapetum at the mononuclear stage, which resulted in pollen dysfunction. The k-means clustering analysis of differentially expressed genes (DEGs) revealed that these genes are related to processes associated with the accumulation of reactive oxygen species (ROS), including electron transport chain, F₁F₀-ATPase proton transport, positive regulation of superoxide dismutase (SOD), hydrogen peroxide catabolic, and oxidation-reduction. Biochemical analysis indicated that ROS-scavenging capability was lower in 722HA than in 722HB, resulting in an accumulation of excess ROS, which is consistent with the transcriptome results. Taken together, these results demonstrate that excessive ROS accumulation may affect the normal development of microspores. Our study provides new insight into the molecular mechanism of pollen abortion in 722HA and will promote further studies of kenaf hybrids.

Male sterility (MS) plays a key role in the hybrid breed production of plants. Researchers have focused on the association between genetic male sterility (GMS) and cytoplasmic male sterility (CMS) in kenaf. In this study, P9BS (a natural GMS mutant of the kenaf line P9B) and male plants of P9B were used as parents in multiple backcross generations to produce P9SA, a CMS line with stable sterility, to explore the molecular mechanisms of the association between GMS and CMS. The anthers of the maintainer (P9B), GMS (P9BS), and CMS (P9SA) lines were compared through phenotypic, cell morphological, physiological, biochemical observations, and transcriptome analysis. Premature degradation of the tapetum was observed at the mononuclear stage in P9BS and P9SA, which also had lower activity of reactive oxygen species (ROS) scavenging enzymes compared with P9B. Many coexpressed differentially expressed genes were related to ROS balance, including ATP synthase, electron chain transfer, and ROS scavenging processes were upregulated in P9B. CMS plants had a higher ROS accumulation than GMS plants. The MDA content in P9SA was 3.2 times that of P9BS, and therefore, a higher degree of abortion occurred in P9SA, which may indicate that the conversion between CMS and GMS is related to intracellular ROS accumulation. Our study adds new insights into the natural transformation of GMS and CMS in plants in general and kenaf in particular.

In order to create transgenic male sterile germplasm in kenaf, the partial length of gene HcPDIL-2a was transformed into maintainer line 722HB via pollen tube pathway. A low-temperature sensitive male sterile mutant called ‘722THS’ was accquired, which showed sterile when grown in winter in Hainan and fertile when grown in summer in Nanning, and a stable Genic Male Sterile (GMS) line 722HS was selected from its sister progenies. A cytoplasmic male sterile line 722HA was bred by crossing the mutant 722THS with the wild type 722B. Cytological observation indicated that the microspore abortion of 722THS and 722HA both happened at dinuclear stage. The mitochondrial DNA molecular tag revealed that mitochondrial DNA rearrangement occurred in 722HA, while mitochondrial DNA of 722THS and 722B remained unchanged. This study provided a new way for the creation of male sterile germplasms, which had important scientific significances and broad application prospects.