Cytoplasmic Male Sterile Lines Research Articles

Transcriptomic and Proteomic Analyses of Celery Cytoplasmic Male Sterile Line and Its Maintainer Line.

Male sterility is a common phenomenon in the plant kingdom and based on the organelles harboring the male-sterility genes, it can be classified into the genic male sterility (GMS) and the cytoplasmic male sterility (CMS). In every generation, CMS can generate 100% male-sterile population, which is very important for the breeders to take advantage of the heterosis and for the seed producers to guarantee the seed purity. Celery is a cross-pollinated plant with the compound umbel type of inflorescence which carries hundreds of small flowers. These characteristics make CMS the only option to produce the commercial hybrid celery seeds. In this study, transcriptomic and proteomic analyses were performed to identify genes and proteins that are associated with celery CMS. A total of 1255 differentially expressed genes (DEGs) and 89 differentially expressed proteins (DEPs) were identified between the CMS and its maintainer line, then 25 genes were found to differentially expressed at both the transcript and protein levels. Ten DEGs involved in the fleece layer and outer pollen wall development were identified by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses, most of which were down-regulated in the sterile line W99A. These DEGs and DEPs were mainly enriched in the pathways of "phenylpropanoid/sporopollenin synthesis/metabolism", "energy metabolism", "redox enzyme activity" and "redox processes". Results obtained in this study laid a foundation for the future investigation of mechanisms of pollen development as well as the reasons for the CMS in celery.

Integrated transcriptomic and proteomic analysis of a cytoplasmic male sterility line and associated maintainer line in soybean.

Heterosis is a critical phenomenon in crop improvement. Cytoplasmic male sterility (CMS) and Restorer gene (Rf) systems are essential components for heterosis-based breeding. However, the molecular mechanism underlying CMS remains largely unclear in soybean. We integrated a morphological investigation with comparative analyses of transcriptomic and proteomic changes in pollen from the CMS line W931A and its maintainer line, W931B, at the uninucleate microspore (UM) and binucleate pollen (BP) stages. Compared to W931B, which had healthy, oval pollen grains, W931A showed shrunken or degraded pollen grains with an irregularly thickened endothelium and decreased starch accumulation. Transcriptomic comparisons revealed a total of 865 differentially expressed genes (DEGs) in W931A over the two stages. These genes were primarily associated with pentose and glucuronate interconversions, sphingolipid metabolism, and glycerolipid metabolism. Proteomic analysis revealed 343 differentially expressed proteins (DEPs), which were mainly involved in carbon metabolism, glycolysis/gluconeogenesis, and nitrogen metabolism. Consistently, Gene Ontology (GO) biological process terms related to pollen development were enriched among DEGs at the UM and BP stages. Notably, four genes with demonstrated roles in pollen development were differentially expressed, including AGAMOUS-LIKE 104, PROTEIN-TYROSINE-PHOSPHATASE 1, and PHOSPHOLIPASE A2. A total of 53 genes and the corresponding proteins were differentially expressed in W931A at both the UM and BP stages, and many of these were pectinesterases, polygalacturonases, peroxidases, and ATPases. The results of this study suggest that pollen development in W931A is likely regulated through suppression of the identified DEGs and DEPs. These findings increase our understanding of the molecular mechanism underlying CMS in soybean, aiding future research into soybean fertility and promoting the efficient use of heterosis for soybean improvement.

Yuan Longping, a great world hunger fighter

Yuan Longping, a great world hunger fighter

Genetic Diversity Analysis and Core Germplasm Collection Construction of Radish Cultivars Based on Structure Variation Markers.

Radish is an economically important root vegetable worldwide. In this study, the 217 cultivated radish accessions were collected and genotyped. To detect the genotypes of these accessions, a total of 24 structure variation (SV) markers distributed on nine chromosomes were employed to analyze genetic diversity and construct a core germplasm collection of radish. The results of polymorphism information content (PIC) indicated a good polymorphism of these SV markers. Population structure analysis and principal component analysis (PCA) results showed that the 217 radish accessions fell into three main populations (P1, P2, and P3). Genetic diversity analysis showed that these populations were highly associated with geographical distribution. The values of the fixation index (FST) indicated a high genetic diversity between P2 and P3, and a moderate genetic diversity between P1 and P2, and P1 and P3. Furthermore, the 43 core germplasm were exploited for creating cytoplasmic male sterility (CMS) lines and cultivating new radish varieties. The high genetic diversity of 217 radish germplasms will not only provide valuable resources for future genetic mapping and functional genomic research, but also facilitate core germplasm utilization and the molecular breeding of radish.

Effect of Gibberellic Acid on Malaysia Cytoplasmic Male Sterile Line During Hybrid Rice Seed Multiplication

Effect of Gibberellic Acid on Malaysia Cytoplasmic Male Sterile Line During Hybrid Rice Seed Multiplication

Population structure and genetic diversity of a germplasm for hybrid breeding in rye (Secale cereale L.) using high-density DArTseq-based silicoDArT and SNP markers

Investigating genetic structure and diversity is crucial for the rye hybrid breeding strategy, leading to improved plant productivity and adaptation. The present study elucidated the population structure and genetic diversity of 188 rye accessions, comprising 94 pollen fertility restoration lines (RF) and 94 cytoplasmic male-sterile (CMS) lines with Pampa sterilizing cytoplasm using SNP and silicoDArT markers from the diversity array technology (DArT)-based sequencing platform (DArTseq). Expected heterozygosity (He) and Shanon’s diversity (I) indexes varied slightly between marker systems and groups of germplasms (He = 0.34, I = 0.51 for RF and CMS lines genotyped using SNPs; He = 0.31, I = 0.48, and He = 0.35, I = 0.53 for RF and CMS using silicoDArTs, respectively). ANOVA indicated moderate variation (7%) between RF and CMS breeding materials. The same parameter varied when chromosome-assigned markers were used and ranged from 5.8% for 5R to 7.4% for 4R. However, when silicoDArT markers were applied, the respective values varied from 6.4% (1R) to 8.2% (3R and 4R). The model-based (Bayesian) population structure analysis based on the total marker pool identified two major subpopulations for the studied rye germplasm. The first one (P1) encompasses 93 RF accessions, and the second one (P2) encompasses 94 CMS and one RF accession. However, a similar analysis related to markers assigned to selected chromosomes failed to put plant materials into any of the populations in the same way as the total marker pool. Furthermore, the differences in grouping depended on marker types used for analysis.

Development of male sterile lines of CMS chilies (Capsicum annuum L.) from F1 hybrids.

Selfing and crossing methods were used to develop the cytoplasmic male sterility (CMS) lines from 2 elite F1 hybrids of CMS hot chilies. The pungency of the CMS lines was improved by backcrossing with the B cultivar. The first and second backcrossed progenies of the CMS lines showed significantly higher capsaicin contents than the F1 hybrids. One good female line K16 × BBC2 (K16), was selected and backcrossed with 3 good maintainer cultivars, C5, C9 and C0. Some incomplete male sterility of pollens was demonstrated in the F1 hybrids and the 1st backcrossed progenies while the partial sterility disappeared by the stage of the second and third generations of backcrossing. When K16 and P32 were crossed with restorers, fruit yields and yield components of certain F1 hybrids, parental lines and commercial varieties were significantly different. Heterosis of yield and yield components of the F1 hybrid chilies was significant. When K16 was used as a female parent, positive and significant heterosis of the F1 hybrids was the same as P32. Moreover, significant GCA of the restorer lines, C7, C8 and C9, was observed in some horticultural characteristics. Furthermore, significant differences of the specific combining ability of some characteristics were observed in a few F1 hybrids.

Genome-Wide Identification and Expression Analysis of TUA and TUB Genes in Wheat (Triticum aestivum L.) during Its Development.

Microtubules play a fundamental role in plant development, morphogenesis, and cytokinesis; they are assembled from heterodimers containing an α-tubulin (TUA) and a β-tubulin (TUB) protein. However, little research has been conducted on the TUA and TUB gene families in hexaploid wheat (Triticum aestivum L.). In this study, we identified 15 TaTUA and 28 TaTUB genes in wheat. Phylogenetic analysis showed that 15 TaTUA genes were divided into two major subfamilies, and 28 TaTUB genes were divided into five major subfamilies. Mostly, there were similar motif compositions and exon-intron structures among the same subfamilies. Segmental duplication of genes (WGD/segmental) is the main process of TaTUA and TaTUB gene family expansion in wheat. It was found that TaTUA and TaTUB genes presented specific temporal and spatial characteristics based on the expression profiles of 17 tissues during wheat development using publicly available RNA-seq data. It was worth noting, via qRT-PCR, that two TaTUA and five TaTUB genes were highly expressed in fertile anthers compared to male sterility. These were quite different between physiological male sterile lines and S-type cytoplasmic male sterile lines at different stages of pollen development. This study offers fundamental information on the TUA and TUB gene families during wheat development and provides new insights for exploring the molecular mechanism of wheat male sterility.

Cyanobacteria Application Ameliorates Floral Traits and Outcrossing Rate in Diverse Rice Cytoplasmic Male Sterile Lines.

In rice, cytoplasmic male sterility (CMS) represents an irreplaceable strategy for producing high-yielding hybrid rice based on the commercial exploitation of heterosis. Thereupon, enhancing floral traits and outcrossing rates in CMS lines increase hybrid seed production and ensure global food security. The exogenous application of cyanobacteria could enhance outcrossing rates in CMS lines and, accordingly, hybrid rice seed production. In the present study, we aimed at exploring the impact of cyanobacteria implementation such as Anabaena oryzae, Nostoc muscorum, and their mixture to promote the floral traits, outcrossing rates, and seed production in hybrid rice. The impact of cyanobacteria (Anabaena Oryza (T2), Nostoc muscorum (T3), and their combination (T4) versus the untreated control (T1) was investigated for two years on the growth, floral, and yield traits of five diverse CMS lines, namely IR69625A (L1), IR58025A (L2), IR70368A (L3), G46A (L4), and K17A(L5). The evaluated CMS lines exhibited significant differences in all measured floral traits (days to heading (DTH), total stigma length (TSL), stigma width (SW), duration of spikelet opening (DSO), spikelet opening angle (SOA)). Additionally, L4 displayed the uppermost total stigma length and stigma width, whereas L1 and L5 recorded the best duration of spikelet opening and spikelet opening angle. Notably, these mentioned CMS lines exhibited the highest plant growth and yield traits, particularly under T4 treatment. Strong positive relationships were distinguished between the duration of the spikelet opening, panicle exertion, panicle weight, seed set, grain yield, total stigma length, spikelet opening angle, stigma width, and number of fertile panicles per hill. Cyanobacteria is a potential promising tool to increase floral traits and seed production in hybrid rice.

The Use of the Polish Germplasm Collection of Nicotiana tabacum in Research and Tobacco Breeding for Disease Resistance

The Polish germplasm collection of Nicotiana tabacum was started in the 1920s. Up to now, more than eight hundred accessions originating from different regions of the world have been gathered in the collection. It includes valuable breeding lines and obsolete cultivars, among them cytoplasmic male-sterile lines. Numerous cultivars are rich sources of features desired in tobacco breeding. Therefore, the accessions are continually characterised in terms of their various features, one of the most important of which is disease resistance. Much research is being done to explain the nature of resistance and its genetic basis. Moreover, cultivars with good agronomic characteristics are used in wide hybridisation, being recipients of resistance genes from wild species or are genetically modified with transgenes conditioning resistance. The biological diversity of cultivars also allows a proper selection of plant material for pathogen studies, while the large number of the accessions facilitates research into the conditions for long seed storage. Numerous examples of the use of Polish tobacco germplasm in research and breeding, specifically in disease resistance, have been presented in this paper.

Comparative Transcriptome Analysis Reveals Key Insights into Fertility Conversion in the Thermo-Sensitive Cytoplasmic Male Sterile Wheat.

Thermo-sensitive cytoplasmic male sterility (TCMS) plays a crucial role in hybrid production and hybrid breeding; however, there are few studies on molecular mechanisms related to anther abortion in the wheat TCMS line. In this study, FA99, a new wheat thermo-sensitive cytoplasmic male sterility line, was investigated. Fertility conversion analysis showed that FA99 was mainly controlled by temperature, and the temperature-sensitive stage was pollen mother cell formation to a uninucleate stage. Further phenotypic identification and paraffin section showed that FA99 was characterized by indehiscent anthers and aborted pollen in a sterile environment and tapetum was degraded prematurely during the tetrad period, which was the critical abortion period of FA99. The contents of O2-, H2O2, MDA and POD were significantly changed in FA99 under a sterile environment by the determination of physiological indexes. Furthermore, through transcriptome analysis, 252 differentially expressed genes were identified, including 218 downregulated and 34 upregulated genes. Based on KOG function classification, GO enrichment and KEGG pathways analysis, it was evident that significant transcriptomic changes in FA99 under different fertility environments, and the major differences were "phenylalanine metabolism", "phenylpropanoid biosynthesis", "cutin, suberine and wax biosynthesis", "phenylalanine, tyrosine and tryptophan biosynthesis" and "citrate cycle (TCA cycle)". Finally, we proposed an intriguing transcriptome-mediated pollen abortion and male sterility network for FA99. These findings provided data on the molecular mechanism of fertility conversion in thermo-sensitive cytoplasmic male sterility wheat.

Evaluation of Advanced Mutant Restorer Lines for Enhancing Outcrossing Rate and Hybrid Seed Production of Diverse Rice Cytoplasmic Male Sterile Lines

Hybrid rice seed production predominantly relies on the pollen ability of restorer lines and the stigma properties of the cytoplasmic male sterile (CMS) lines. Improving the pollen ability and agronomic performance of restorer lines could be achieved using mutation to reinforce the outcrossing rate and seed set percentage of CMS lines. Two commercial restorer cultivars (Giza-178 and Giza-179) were treated with three different doses of gamma-ray; 200, 300, and 400 Gy. The derived mutant restorer lines were selfed and constantly evaluated with their corresponding parental cultivars until the M6 generation. Six promising mutant lines were chosen based on their performance to be evaluated with their parents for their pollen and agronomic characteristics. The novel-induced mutant restorer lines exhibited significant differences in all studied pollen and agronomic characteristics. The mutant restorer lines R4 and R7 exhibited superior desirable anther length (2.36 and 2.38 mm, in the same order), anther width (0.47 and 0.45 mm), pollen fertility (97.50 and 97.31%), basal pore length (218.8 and 299.5 mm), apical length (103.6 and 108.1), number of pollen grains per anther (1810 and 1832), plant height (121.8 and 115.9 cm), fertile tillers (23.50 and 18.89), number of spikelets per panicle (203.2 and 202.5), panicle length (25.34 and 24.26 cm), number of filled grains per panicle (195.5 and 191.8), seed set percentage (95.56 and 96.63%), and grain yield (11.01 and 11.36 t/ha). Based on the results of the first two seasons, the two novel promising restorers and their parental cultivars were crossed with four diverse CMS lines. The seed set percentage and hybrid seed production of the crossed CMS lines were evaluated for further two seasons. The results exposed that the combinations derived from restorer lines R4 and R7 with most of the evaluated CMS lines produced the uppermost seed set percentage outcrossing rate. Particularly in combination with CMS2, the restorer lines R4 and R7 displayed the highest seed set percentage (39.57 and 34.80%), number of tillers fertile per hill (25.31 and 23.32), seed yield (2.02 and 1.81 t/ha), and harvest index (23.88 and 22.66%). Conclusively, the derived gamma-ray-induced mutant restorer lines R4 and R7 could be exploited as a new source for ensuring desirable pollen and anther characteristics in order to improve the outcrossing rate of CMS lines and hybrid rice seed production.

Gw2.1, a new allele of GW2, improves grain weight and grain yield in rice

Grain weight is an important characteristic of grain shape and a key contributing factor to the grain yield in rice. Here, we report that gw2.1, a new allele of the Grain Width and Weight 2 (GW2) gene, regulates grain size and grain weight. A single nucleotide substitution in the coding sequence (CDS) of gw2.1 resulted in the change of glutamate to lysine (E128K) in GW2.1 protein. Complementation tests and GW2 overexpression experiments demonstrated that the missense mutation in gw2.1 was responsible for the phenotype of enlarged grain size in the mutant line jf42. The large grain trait of the near-isogenic line NIL-gw2.1 was found to result from increased cell proliferation during flower development. Meanwhile, NIL-gw2.1 was shown to increase grain yield without compromising the grain quality. The GW2 protein was localized to the cell nucleus and membrane, and interacted with CHB705, a subunit of the chromatin remodeling complex. Finally, the F1 hybrids from crosses of NIL-gw2.1 with 7 cytoplasmic male-sterile lines exhibited large grains and desirable grain appearance. Thus, gw2.1 is a promising allele that could be applied to improve grain yield and grain appearance in rice. Availability of data and materialsThe datasets generated and/or analyzed in the study are available from the corresponding author on reasonable request.

QTL Analysis and Heterosis Loci of Effective Tiller Using Three Genetic Populations Derived from Indica-Japonica Crosses in Rice

Effective panicle numbers (PNs) and Tiller numbers (TNs) are important traits affecting rice (Oryza sativa L.) architecture and grain yield. However, the molecular mechanisms underlying PN and TN heterosis remain unknown in rice. In addition, new PN- or TN-related genes need to be detected and discovered. In this study, in order to detect rice quantitative trait loci (QTLs) and the heterosis-related loci of PN or TN in rice, we developed a high generation recombinant inbred line (RIL) population from a cross of two elite cultivars, Luohui9 (Xian/Indica) and RPY geng (Geng/Japonica), and two testcross hybrid populations derived from the crosses of RILs and two cytoplasmic male sterile lines, YTA (Xian/Indica) and Z7A (Geng/Japonica). Finally, nine QTLs of PN across four seasons were identified, and two QTLs of TN in 191HB were mapped. Besides this, six heterosis-related QTLs of PN and five heterosis-related QTLs of TN were located. We found that heterosis-related QTLs of PN or TN covered multiple known genes, such as MOC1, TAC1 and OsETR2. Furthermore, homologous gene analysis identified one candidate gene of PN (LOC_10g25720). Together, these findings uncover multiple heterosis-related loci, and provide a new insight into the heterosis mechanism of PN and TN in rice.

Increasing seed set and pollen proofing in Brassica juncea and Brassica napus through novel non-woven synthetic pollination control bags

ABSTRACT Brassica juncea and B. napus species are predominantly self-pollinated. Parchment paper or muslin cloth bags are used for controlled selfed or cross-seed production. We evaluated three non-woven synthetic pollination control bags (PCBs) for two years as replacement for the commonly used PCBs. The new synthetic PCBs were fully pollen proof since the molecular marker analysis of the random seeds set on two cytoplasmic male sterile (CMS) lines isolated with PCBs were all maternally originated. DWB03 bags returned 47% significantly more seeds per bud than the parchment paper bags. Larger DWB03 bags showed 38% higher mean seed yield (g/plant) than the muslin cloth bags. Re-used DWB03 bags were statistically on par with new bags for the seed yield per plant. On average, DWB03 bags produced 57% more hybrid seed set on CMS lines than the muslin cloth bags. The micro-climate in DWB03 bags appeared to be more conducive for good seed set as compared to the outside and muslin cloth bags. Non-woven PCBs offer better options for replacing paper or muslin cloth bags in the breeding of Brassicas.

De novo assembly of two chromosome-level rice genomes and bin-based QTL mapping reveal genetic diversity of grain weight trait in rice.

Following the “green revolution,” indica and japonica hybrid breeding has been recognized as a new breakthrough in further improving rice yields. However, heterosis-related grain weight QTLs and the basis of yield advantage among subspecies has not been well elucidated. We herein de novo assembled the chromosome level genomes of an indica/xian rice (Luohui 9) and a japonica/geng rice (RPY geng) and found that gene number differences and structural variations between these two genomes contribute to the differences in agronomic traits and also provide two different favorable allele pools to produce better derived recombinant inbred lines (RILs). In addition, we generated a high-generation (> F15) population of 272 RILs from the cross between Luohui 9 and RPY geng and two testcross hybrid populations derived from the crosses of RILs and two cytoplasmic male sterile lines (YTA, indica and Z7A, japonica). Based on three derived populations, we totally identified eight 1,000-grain weight (KGW) QTLs and eight KGW heterosis loci. Of QTLs, qKGW-6.1 and qKGW-8.1 were accepted as novel KGW QTLs that have not been reported previously. Interestingly, allele genotyping results revealed that heading date related gene (Ghd8) in qKGW-8.1 and qLH-KGW-8.1, can affect grain weight in RILs and rice core accessions and may also play an important role in grain weight heterosis. Our results provided two high-quality genomes and novel gene editing targets for grain weight for future rice yield improvement project.

Comparison of Mitochondrial Genomes between a Cytoplasmic Male-Sterile Line and Its Restorer Line for Identifying Candidate CMS Genes in Gossypium hirsutum.

As the core of heterosis utilization, cytoplasmic male sterility (CMS) has been widely used in hybrid seed production. Previous studies have shown that CMS is always closely related to the altered programming of mitochondrial genes. To explore candidate CMS genes in cotton (Gossypium hirsutum), sequencing and de novo assembly were performed on the mitochondrial genome of the G. hirsutum CMS line SI3A, with G. harknessii CMS-D2 cytoplasm, and the corresponding G. hirsutum restorer line 0-613-2R. Remarkable variations in genome structure and gene transcripts were detected. The mitochondrial genome of SI3A has three circle molecules, including one main circle and two sub-circles, while 0-613-2R only has one. RNA-seq and RT-qPCR analysis proved that orf606a and orf109a, which have a chimeric structure and transmembrane domain, were highly expressed in abortive anthers of SI3A. In addition, comparative analysis of RNA-seq and full-length transcripts revealed the complex I gene nad4 to be expressed at a lower level in SI3A than in its restorer and that it featured an intron retention splicing pattern. These two novel chimeric ORFs and nad4 are potential candidates that confer CMS character in SI3A. This study provides new insight into the molecular basis of the nuclear–cytoplasmic interaction mechanism, and that putative CMS genes might be important sources for future precise design cross-breeding of cotton.

TaEXPB5 functions as a gene related to pollen development in thermo-sensitive male-sterility wheat with Aegilops kotschyi cytoplasm

The thermo-sensitive cytoplasmic male-sterility line with Aegilops kotschyi cytoplasm (K-TCMS) is completely male sterile under low temperature (< 18 ℃) during Zadoks growth stages 45–52, whereas its fertility can be restored under hot temperature (≥ 20 ℃). The K-TCMS line may facilitate hybrid breeding and hybrid wheat production. Therefore, to elucidate the molecular mechanisms of its male sterility/fertility conversion, we conducted the association analysis of proteins and transcript expression to screen fertility related genes using RNA-seq, iTRAQ, and PRM-based assay. A gene encoding expansin protein in wheat, TaEXPB5, was isolated in K-TCMS line KTM3315A, which upregulated expression in the fertility anthers. Subcellular localization analysis suggested that TaEXPB5 protein localized to nucleus and cell wall. The silencing of TaEXPB5 displayed pollen abortion and the declination of fertility. Further, cytological investigation indicated that the silencing of TaEXPB5 induced the early degradation of tapetum and abnormal development of pollen wall. These results implied that TaEXPB5 may be essential for anther or pollen development and male fertility of KTM3315A. These findings provide a novel insight into molecular mechanism of fertility conversion for thermo-sensitive cytoplasmic male-sterility wheat, and contribute to the molecular breeding of hybrid wheat in the future.

Progress in identifying male sterility genes and utilizing heterosis in wheat

<p indent="0mm">Male sterility is a valuable trait for plant breeding and heterosis utilization. Identifying and characterizing male-sterile genes in crops have improved our understanding of the molecular mechanisms controlling anther and pollen grain development, and enabled the development and effective use of many biotechnology-based male sterile systems in hybrid crop breeding. Research progress on male-sterile genes in wheat (<italic>Triticum</italic> spp.) lags that in rice (<italic>Oryza sativa</italic>) and maize (<italic>Zea mays</italic>) because of the polyploid nature and complex genome of wheat; therefore, only a few wheat male-sterile genes have been identified or cloned. Due to the limited progress in understanding the basis of male sterility, a low-cost and efficient wheat hybridization system has not been developed. In this review, we summarize the progress in characterizing genetic male sterility (GMS), cytoplasmic male sterility (CMS), and environment-sensitive GMS (EGMS) in wheat. At least five GMS genes were reported in wheat via forward genetic approaches, of which the dominant gene <italic>Ms2</italic> and the recessive genes <italic>ms1</italic> and <italic>ms5</italic> have been cloned. Furthermore, clustered regularly interspaced short palindromic repeats (CRISPR)/CRISPR-associated protein (Cas) gene editing technology was used to characterize the role of four wheat homologs of maize <italic>Ms26</italic> and <italic>Ms45</italic>, rice <italic>NO POLLEN1 </italic>(<italic>OsNP1</italic>), and <italic>Arabidopsis thaliana</italic> <italic>DEFECTIVE IN TAPETAL DEVELOPMENT AND FUNCTION 1 </italic>(<italic>TDF1</italic>) in male fertility. Over 70 different types of wheat CMS systems have been reported; in most cases, CMS is caused by mitochondrial genome rearrangements, and can be reversed with nuclear restorer of fertility (<italic>Rf</italic>) genes. Current approaches to identify wheat <italic>Rf</italic> genes for fertility restoration focus on T-type (<italic>Triticum timopheevii</italic>) and K-type (<italic>Aegilops</italic><italic>kotschyi</italic> Boiss) CMS lines. At least 15 wheat <italic>Rf</italic> genes have been identified, but only two,<italic> Rf1</italic> and <italic>Rf3</italic>, were cloned by genetic mapping and comparative sequence analysis. These two restorer genes and their mode of action in CMS provide new opportunities for wheat hybrid breeding. The candidate CMS restorer genes <italic>Rfk1</italic> and <italic>Rf</italic> ^{<italic>multi</italic>} were also predicted using comparative genomics approaches. EGMS may involve epistatic regulation of non-coding RNAs that restore fertility under specific conditions, and thus provides a special class of germplasm for hybrid production in several crops; however, only a few wheat EGMS genes have been reported and none of them have been cloned. Next, we summarize the wheat hybridization systems that rely on male-sterile genes, such as the “three-line system” based on CMS genes, the “two-line system” based on EGMS genes, and the next generation of hybrid seed production based on nuclear GMS genes. Recent efforts in hybrid wheat breeding have produced hybrid varieties with desirable attributes, which have been marketed in Europe and China. Finally, we discuss future perspectives and challenges in the research and application of next-generation wheat hybrid production technology. This review provides a reference for researching and utilizing male-sterile genes in <italic>Triticeae</italic> crops.

Fine mapping of Rf2, a minor Restorer-of-fertility (Rf) gene for cytoplasmic male sterility in chili pepper G164 (Capsicum annuum L.).

Genome re-sequencing and recombination analyses identified Capana06g000193 as a strong candidate for the minor male fertility restoration locus Rf2 in chili pepper G164 harboring two dominant male fertility restoration genes. Male fertility restoration genes of chili pepper restorer line G164 (Capsicum annuum L.) were studied using molecular marker genotypes of an F2 population (7G) of G164 crossed with the cytoplasmic male sterility line 77013A. The ratio of sterile to fertile single plants in the F2 population was 1:15. This result indicates that chili pepper G164 has two dominant restoration genes, which we designated as Rf1 and Rf2. An individual plant recessive for Rf1 and heterozygous for Rf2, 7G-112 (rf1rf1Rf2rf2), was identified by molecular marker selection and genetic analysis, and a single Rf2 gene-segregating population with a 3:1 ratio of fertile to sterile plants was developed from the self-pollination of male fertile individuals of 77013A and 7G-112 hybrid progeny. Bulk segregant analysis of fertile and sterile pools from the segregating populations was used to genetically map Rf2 to a 3.1-Mb region on chromosome 6. Rf2 was further narrowed to a 179.3-kb interval through recombination analysis of molecular markers and obtained the most likely candidate gene, Capana06g000193.