Comparative transcriptome analysis provides insights into the gene regulation network of cytoplasmic male sterility in chilli pepper

Rice is a strictly self-pollinating crop. However, in hybrid rice seed production, an effective male sterility system is used to produce hybrid seed in bulk. In hybrid rice system, the pollen grains of cytoplasmic male sterile (CMS) are sterile and the female organ of the CMS depends on the fertile pollen released by the maintainer or restorer lines via out-crossing or cross-pollination in order to produce seed. Floral trait and flowering behavior of CMS and its corresponding maintainer or restorer lines are essential factors in hybrid rice seed production because they influenced the out-crossing or cross-pollination between parental lines. Two local CMSs and their corresponding maintainer lines were developed through breeding program in Malaysian Agricultural Research and Development Institute (MARDI) namely 0025A/0025B and 0047A/0047B. This study was carried out on floral traits and flowering behavior of these two hybrid line. Present studies have shown that there were variations between the CMS and its maintainer lines whether on floral trait or flowering behavior for both hybrid rice combinations. The results showed that stigma characters for both 0025A and 0047 were superior than their respective maintainers. Therefore, it is expected that the out-crossing rate would be high. Seeding date intervals need to be done on 0025A/0025B during nursery stage because the on-set of flowering between parental lines was significantly different. Panicle of both CMS was also classified as just exserted and partially-exserted and application of exogenous hormones such as gibberellic acid was useful to improve panicle elongation and consequently increase the seed set and yield. Correlation study indicates that the stigma area of both 0025A and 0047A has significant positive correlation with out-crossing rate.

Yuan Longping, is known as ‘Father of Hybrid Rice’, was born into an intellectual family in Beijing on 13 August 1929 and passed away on 22 May 2021, in Changsha, Hunan Province, China. He was an academician of the Chinese Academy of Engineering, a foreign associate of the National Academy of Sciences of the United States and former director general of China National Hybrid Rice Research & Development Center. He was a person who had been more passionate about his job development of hybrid rice for food security in the world and was full of the sense of mission and responsibility. This builds up his prominent scientific spirit: persistent pursuit and innovating forever. When Yuan was a boy, his teacher once took the pupils to go on an excursion. In a horticulture garden, he saw green grass, fresh flowers, red peaches and bunches of grapes. The lively energy in the great nature gave him a deep impression in his young mind. At that time, an American comic movie ‘Modern Times’ described the countryside as a beautiful heaven and Yuan was attracted to the movie. The superposition of the above two motivated him to study agriculture. In 1949, Yuan completed his high school courses and enrolled at the Southwestern Agricultural College in Chongqing, majoring in agronomy. This marked the beginning of his lifelong work in agriculture. After his graduation from college in 1953, Yuan did a teaching job at An-jiang Agricultural School in An-jiang county in the western mountainous area of Hunan, China, and married Deng Ze in 1964. As a teacher, he taught botany, crop cultivation, breeding and genetics and even Russian language. Therefore, while teaching, he conducted scientific experiments to improve food crop production, involving asexual crosses between crops, using Michurin's theory. However, after 3 years of research, he found that the theory was incorrect, so he gave up asexual crossing experiments and recognized Mendel's modern genetics. Just at that time, he happened to discover a ‘Natural rice hybrid’. However, in the early 1960s, no one thought that hybrid vigour can be exploited in a self-pollinated crop such as rice and no solutions for seed production of high-yielding hybrid in self-pollinated crop. But Yuan believed that heterosis was a universal phenomenon, and rice was no exception. The ‘Natural rice hybrid’ enlightened his courage to challenge classical theory. He made up his mind to research hybrid rice and started to explore the mechanism of heterosis utilization in rice after the discovery of this extraordinary hybrid in 1964. As widely known, heterosis is best expressed in the first-generation hybrid of two genetically distinct parents. Rice is a self-pollinated crop. Its flower is tiny, and one flower can only produce one grain. These characteristics make it impossible to remove pollen-producing anthers manually and thus severely prevent the commercial application of hybrid in rice before the 1970s. Yuan discovered the male sterile rice plants in 1964 and 1965 and then published his first research paper on rice male sterility in the Chinese Science Bulletin, which initiated China's hybrid rice researches in 1966. In this landmark paper, he rightfully predicted that rice heterosis could be utilized to increase yield through the development of male sterile (or A) lines, maintainer (or B) lines and male restore (or R) lines and described the nice blueprint of utilizing rice heterosis successfully. Because Yuan insisted on his research on developing hybrid rice in the late 1960s, he moved to Hainan and Yunnan from Hunan in the winter like a migratory bird. It was on Hainan Island in 1970 that a male sterile plant in wild rice—‘WA’ was found. This nature wild male-abort rice provided a crucial genetic tool in the birth of China's commercial hybrid rice and confirmed Pasteur's famous aphorism ‘Chance favors the prepared mind only’. This led to the promising discovery progress in the development of hybrid rice. After 9 years of hard work, Yuan and his research team successfully developed the three genetic lines in 1973, that is, male sterile line, maintaining line and restore line or A, B and R line, which are essential for developing hybrid rice. In 1974, he selected the first hybrid variety Nanyou No.2, which showed very strong heterosis and high yield potential. In 1975, technology for commercial hybrid seeds production was completed. One year later, hybrid rice was released for commercial production. Thus, Yuan is considered as the first scientist to successfully alter the self-pollinated characteristics of rice and facilitate the large-scale production of hybrid rice, which has 20% more yield than elite inbred varieties. Dr. M.S. Swaminathan, the former director general of the International Rice Research Institute (IRRI) gave a speech at an international rice conference at IRRI in 1982: ‘Yuan Longping, we called him the father of hybrid rice, is well-deserved, and his success is not only the pride of China, but the pride of the world, his achievement has brought the gospel to the world’. He indicated in his message to Yuan on the occasion of Yuan's 80th birthday: ‘You took the unknown path into an area full of skepticism and made the impossible possible-achieve the rice hybrid miracle’. Yuan had a dream called the dream under the rice panicle. Once, he dreamed that the rice plants were as high as a sorghum plant; the panicles were as large as besoms; and the grains were as big as peanuts. Yuan and his assistants were enjoying the shade under the rice panicles. Actually, it is a dream of rice high yield. Indeed, Yuan was a person who was not satisfied with his past success. He continued dreaming of pursuing the dream of rice high yield. After achieving the success of the three-line (A, B and R) hybrid rice, he kept on exploring and had always been thinking about how to develop new approaches to enhance the heterosis level and simplify the methodology for hybrid rice breeding. In 1986, Yuan proposed the ‘three-step’ strategy of developing hybrid rice. He was a responsibility expert for the programme of developing two-line hybrid rice also. By composition of photo-thermo-sensitive genic male sterile (PTGMS) and restorer line (R), the two-line system (PTGMS, R) is simpler than the three-line system (A, B and R), that is, a system that would not require a maintainer line (B), or the PTGMS can plan as both the sterile line (A) and the maintainer line (B), because the PTGMS on one hand shows sterility under the condition of high temperature and long sunshine and on the other hand returns to normal fertility under the condition of flat temperature and short sunshine. So, people can use the PTGMS as sterile line (A) to produce the hybrid rice seeds in hot summer and reproduce PTGMS itself (as maintainer line, B) when the temperature is relatively low and the sunshine is relatively short in spring and autumn. Under Yuan's wise leadership and technical guidance, his research team succeeded in developing a whole package of two-line hybrid rice technology in 1995. Moreover, in the new 21st century, the world faces risks that rice production fails to meet the demand of population increase and planting area recession. Yuan assumed the responsibility to develop super hybrid rice. He laid out a road map: a combination of morphological improvement, inter-subspecies hybridization and foreign genes for high yield introduction with molecular biotechnology. Under the guidance of this road map, his team fulfilled the first, second, third and fourth stage yield goals in a large planting area in the year of 2000 (10.5 t/ha), 2004 (12 t/ha), 2012 (13.5 t/ha) and 2014 (15 t/ha), respectively. To exceed the target of 18 t/ha super hybrid rice programme, he put forward a new strategy again. That was to raise the height of the rice plant on the base of lodging tolerance. He considered that it would help increase rice yield potential. Yuan used to say ‘To pursue crops' high yield is an eternal subject’. The important progresses of new super hybrid rice varieties' demonstration have been made continuously because the above targets were achieved. A new hybrid, Xiangliangyou 900, yielded more than 16 t/ha in 2015, 2016 and 2017 at a 6.8 ha of demonstration in Gejiu, Yunnan. More excitingly, an average yield of 17.28 t/ha for large-scale rice cultivation was achieved at the same demonstration location in 2018. It was the highest yield level in large-scale cultivation in the world. In 2022, a new yield record has been created by variety Zhuoliangyou 1126 with the yield of 18 t/ha on hectare level in the Gejiu demonstration location again. Besides, Yuan's team launched the breeding research of saline-alkali-tolerant hybrid rice, aiming at the application in about 1 billion ha of saline-alkali land in China. The team also carried out the third-generation hybrid rice research, facilitating the integration of freedom of choice of parents for hybrids in two-line system with stable sterility of male sterile line to develop elite hybrids. The representative variety for the third-generation hybrid ‘Sanyou No.1’ has been planted as a late one for double rice a year in Hunan Province since 2019. Its growth period was 125 days, and the yield reached more than 13.5 t/ha. It has made good results in Hengnan County, Hunan Province, and the total yield of double-cropping rice within a year reached 22.96 t/ha in 2020. With Yuan's relentless pursuit and innovation, his research resulted in the realization of increasing rice yield potential. And also, the utilization of rice heterosis has offered an effective way for large-scale improvement in per unit output on current condition. Now, the planting area of hybrid rice expanded greatly, reaching around 1.67 × 107 hm2 in recent years and accounting more than 50% of a total one rice-growing area in China. In recent years, the average yield of rice is approximately 6.45 t/hm2, in which the average yield of hybrid rice is 7.5 t/hm2 and that of conventional rice is 6.15 t/hm2 in China. The two-line hybrid rice is planted 3 million ha annually in China and not only outyielded the three-line hybrids by about 10% but also is popular with good quality. Hybrid rice has saved thousands of lives in a heavily populous China with limited acreage of farming land in the last 30 years. This remarkable success has helped transform China from food deficiency to food security within three decades despite the significant increase in its population. The development of hybrid rice helps China solve food shortage problem successfully and provides additional income to thousands of farmers today. Because of its great success, hybrid rice is honoured as the second ‘Green Revolution’. Yuan's lifelong pursuit was ‘Developing hybrid rice for the welfare of the people all over the world’. Acting on his belief that science should have no boundaries between countries, he shared his knowledge, experience, ideas and valuable breeding materials with scientists outside China to enthusiastically extend hybrid rice technology to other countries. In early 1980s, he donated the indispensable and unique cytoplasmic male sterile (CMS) line to the IRRI; many countries obtained this precious resource through IRRI to develop new male sterile lines and hybrid varieties. Those hybrid rice varieties are now widely planted in many Asian countries. From then on, Yuan visited IRRI cooperative research and technology exchange for more than 30 times. In 1980, China's hybrid rice was transferred to the United States as the first agricultural patent technology. Yuan's research institute had established cooperation relations with the RiceTec Inc., an American seed company, since 1992. Yuan had been to the United States for more than 10 times to guide the research and production of hybrid rice there. He was appointed as the chief consultant of the Food and Agriculture Organization (FAO) of the United Nations and travelled to India, Vietnam, Bangladesh and the Philippines many times. He believed that by taking advantage of China's world leading expertise on technologies and successful experience on hybrid rice research and application, more experts would be able to be trained for the promotion and application of hybrid rice technologies. Up to now, Yuan's research institute and teams already trained over 14,000 scientists or breeders from more than 80 countries. Through such training, Yuan not only imparted his knowledge and wisdom to the trainees but also played as an envoy and erected the bridge of better consolidation of the friendship among all of the trainees and external development of hybrid rice in Asian, African and South American countries. Yuan's concerns went beyond China's food supply and extended to the enormous problem of world hunger. As a result, through Yuan's efforts to extend hybrid rice technology to other countries, hybrid rice have been commercialized spreading throughout Asia and Africa and the Americas including India, Vietnam, Bangladesh, Pakistan, Philippines, Brazil, Madagascar and the United States, where hybrid rice occupied over 8 million ha in 2020. Farmers in more than 10 other countries besides China have thus benefited from his excellent work. The famous American Economist Professor Don Paarlberg commented on his book of Toward a Well-fed World: ‘It illustrates the utility of agricultural science, the caprice of chance, and the accomplishment of a dedicated man … Yuan Longping has bought China valuable time with which to bring down the rate of population growth. As agricultural science advances, the threat of famine retreats. Yuan led toward a well-fed world’. In 2019, for the celebration of the 70th anniversary of the founding of the People's Republic of China, the ‘Medals of the Republic’ were awarded to eight people, including Yuan Longping, in grand recognition of exemplary meritorious figures who have made outstanding contributions to the construction and development of New China and in the promotion of the national spirit and the spirit of the times. In recognition of Yuan's work, he had been bestowed a number of awards and honours also, which includes four of the top prizes in China: the Special-class National Invention Prize in 1981, which was the first of the country; the first National Supreme Scientific and Technology Award in 2001; the Special-class National Science and Technology Award in 2014; and about 20 awards in the world, including the 1987 UNESCO Science Prize, the 1995 FAO World Food Security and Sustainability Medal, the 2004 World Food Prize in the United States and the 2004 Wolf Prize in Israel, and also in 2006, he was elected as a foreign associate of the National Academy of Sciences of the United States. Yuan published more than 70 articles and six monographs on hybrid rice research and development. The book Hybrid Rice Breeding and Cultivation has been recognized as an authoritative reference on hybrid rice technology within China, and two others, A Concise Course on Hybrid Rice and Technology of Hybrid Rice Production, are considered to be primers for hybrid rice technology both within and outside China. Especially, the book, Technology of Hybrid Rice Production, was published by FAO in 1995 and has been translated into other languages. It is considered as the best instruction book in international training of hybrid rice. In 1999, the International Commission on Asteroid Nomenclature named the 8117 asteroid as Yuan Longping Asteroid. Now, Yuan Longping has really been transformed into Yuan Longping Asteroid, hanging high in the sky, overlooking the earth. Yuan Longping's lifelong commitment to putting an end to world hunger has brought mankind one step closer to the full realization of his inborn rights. Along the road, our world has become more peaceful and human beings more humane and decent.

Conversion of GMS male sterility system to CMS will increase the male sterility proportion of female parent of F1 seed production from 50 to 100%. It can eliminate the cost of emasculation and ensure seed hybridity through avoidance of self pollination. In this study, segregation analysis of four male sterile mutants consisting of one CMS mutant (CA1) and three GMS mutants (GA1, GA3 and GA4) showed that each had single recessive gene inheritance. A modified complementation test was performed by replacing male sterile mutants with their maintainer line as male parent. The observation on CMS and GMS male sterility loci revealed that GA1 and GA3 had mutated in both nuclear restorer genes for CMS and GMS, while CA1 and GA4 each carried mutations in single male sterility system of nuclear restorer genes for CMS and GMS, respectively. The important finding is the broader function of a maintainer line in certain male sterility system that can be used as a maintainer or restorer line for other male sterility systems and CC1 is confirmed as the general restorer for both CMS and GMS systems. Bell pepper lacks the CMS nuclear restorer allele Rf and also contains mutations in its GMS nuclear restorer locus. Consequently the GMS maintainer and restorer lines of bell pepper are also the CMS maintainer line. This study has demonstrated the conversion of the male sterility system of the non pungent GMS bell pepper restorer line, GC3, to a CMS male sterile line using S-type cytoplasm donor (CA1) and GC3 has converted to be CMS restorer line using a Rf-allele donor (CC1). Both donors S-type cytoplasm and the Rf allele are derived from tropical pungent pepper that stable on male sterility and fertility restoration, respectively. Evaluation of S-type cytoplasm and Rf-allele introgression was conducted in BC1F1 and BC1F2 populations. Two breeding lines were selected as CMS male sterile candidates and four breeding lines were selected as CMS restorer line candidates of bell pepper. Several morphological traits of the iv selected progeny were measured and shown similar to their recurrent parent (GC3). The effectiveness of molecular markers linked to traits of interest of Stype cytoplasm, pungency and CMS nuclear restorer fertility locus were evaluated in 30 BC1F1 and 116 BC1F2 individuals. Since this study employed the stable of Rf allele and S-type cytoplasm donor from tropical pungent pepper, the developed CMS male sterile line and the restorer line candidates of bell pepper should be stable. Conversion of the male sterility system of bell pepper from GMS to CMS was performed successfully.

Inhibition of chalcone synthase Expression in Anthers of Raphanus sativus with Ogura Male Sterile Cytoplasm

The main objective of the present study was to identify mitochondrial DNA based marker, which can distinguish male sterile and fertile counterparts of the cytoplasmic male sterile (CMS) lines used in production of rice hybrids. Amplified fragment length polymorphism (AFLP) analysis in CMS lines: IR58025A & IR62829A and their respective maintainers: IR58025B & IR62829B identified a polymorphic DNA fragment of about 510 bp size that was present in both CMS (A) and absent in their maintainer (B) lines. Sequencing followed by database analysis of the polymorphic fragment indicated about 97% similarity with mitochondrial NADH gene subunits of rice, maize and wheat. Based on the variable sequence regions, a site specific primer pair (BF-STS-401) was designed. PCR analysis showed that BF-STS-401 could amplify a strong band of 464 bp size in CMS and a faint band of the same size in maintainer line. To act as a positive control and avoid possible errors in PCR, BF-STS-401 was multiplexed with a new primer pair (BF-STS-402), derived from mitochondrial atp9 subunit of rice, producing monomorphic amplification indiscriminately in both CMS and maintainer lines. Both the primer pairs in combination clearly differentiated CMS lines from their corresponding maintainer lines. This primer combination was validated in a set of diverse genotypes consisting of different sources of CMS lines, restorer lines, hybrids, varieties and mixed samples from private seed companies. Our results suggested that the multiplex primer pairs developed in this study can be effectively utilized to assess the genetic purity in commercial seed lots of CMS lines and hybrids of rice.

One-hundred-and-eighty-one nuclear DNA probes were used to examine restriction-fragment length polymorphism in inbred lines of the cultivated sunflower (Helianthus annuus L.). The probes were from six libraries: two genomic libraries - one made with PstI and the other with HindIII, and four cDNA libraries - from etiolated plantlets, green leaves, ovaries, petals and anthers. Total DNA from 17 inbred lines representing an overview of the genetic stocks of sunflower, including restorer and maintainer lines of the classical cytoplasmic male sterility, was digested with four different restriction enzymes and probed in 331 probe-enzyme combinations. Of 181 clones analysed, 73 probes were found to be polymorphic. Genetic distances between inbreds were calculated from the resultant proportion of shared bands and submitted to principal component analysis and the UPGMA 'tree-making' method. The RFLP analysis allowed a clear differentiation between restorer and maintainer lines of the cytoplasmic male sterility, together with a grouping of some of the genotypes from the same origin. The analysis of the accuracy of distance estimation as a function of the number of probe-enzyme combinations used, indicates that 40-50 combinations ensure a confidence level of near 95%. Considering the inbreds as representatives of the range of cultivated inbreds, estimates of gene diversity, as well as estimates of average gene diversity between and within the sets of restorer and maintainer lines, were calculated. Estimation of gene diversity showed that the available genetic variability in cultivated sunflower, based on allelic frequencies, is lower than that of other plants (H=0.20). Moreover, we show that the proportion of genetic variability due to the difference between maintainer and restorer lines (Dm) is about 2%.

Pulsatilla chinensis (Bunge) Regel has been widely used in the pharmaceutical industry. With the deepening of clinical application, the research on its plant resources has attracted much attention. However, the underlying molecular mechanisms of distinct germination during Pulsatilla seed development are still mostly unknown. Therefore, in this study, four germination stages of P. chinensis seeds, with obvious differences in seed appearance traits, were used as materials. Transcriptome sequencing technology was used to analyse the molecular mechanisms of seed germination. A total of 27,601 differentially expressed genes (DEGs) (six different groups) were determined. KEGG enrichment analysis revealed that the up-regulated DEGs were enriched in phenylpropanoid biosynthesis, photosynthesis, photosynthesis–antenna proteins, plant hormone signal transduction, flavonoid biosynthesis and other pathways. A total of 87 DEGs was enriched in phytohormone signal transduction pathways, including auxin (25), abscisic acid (13), gibberellin (6), ethylene (9) and cytokinin (7). Furthermore, a protein–protein interaction network was constructed using these DEGs. Some DEGs were validated by qRT-PCR analysis. This comprehensive analysis provided basic information on the key genes of plant hormone signal transduction pathways involved in the seed germination process of P. chinensis (Bunge) Regel.

Cytoplasmic male sterility (CMS) is a maternally inherited trait that is widely utilized in hybrid seed production. In this study, the Rs-Rf 1 gene and its allele, the Rs-rf 1 gene, were isolated from a radish CMS restorer line and maintainer, respectively. The Rs-Rf 1 gene was identified as a member of the pentatricopeptide repeat family. The results of reverse transcription (RT)-PCR and quantitative RT-PCR analyses of the floral buds at different development stages were used to characterize the expression features of Rs-Rf 1 and orf138, a unique transcribed gene in the mitochondrion for its role in determining male sterility in the cytoplasm. orf138 was up-regulated in the CMS line and F1 hybrid at three bud developmental stages, including meiosis and the tetroid and microspore stages, but it was not expressed in the maintainer and restorer line, suggesting that the Rs-Rf 1 gene was not able to block orf138 transcription. As in the F1 plant, the transcription of Rs-Rf 1 was gradually up-regulated in the restorer line with progressive stages of bud development, but it was not transcribed in the CMS line and maintainer. Different restriction sites for SspI between the allele of Rs-Rf 1 and Rs-rf 1 were also identified, and a guideline for using the functional marker to distinguish the genotypes of individuals with Rs-Rf 1 was developed. Due to its co-dominant character, this marker could be used to discriminate the heterozygous and homozygous fertility-restoring allele. Using this functional marker, we established a system of molecular marker-assisted development of elite CMS lines in radish. This system will contribute greatly to candidate maintainer identification and accelerate the process of elite CMS line development in late-bolting radish breeding programs.

The practice of hybridization has greatly contributed to the increase in crop productivity. A major component that exploits heterosis in crops is the cytoplasmic male sterility (CMS)/nucleus-controlled fertility restoration (Rf) system. The development and use of hybrid rice varieties on commercial scale utilizing male sterility and fertility restoration system has proved to be one of the mile stones in the history of rice improvement. Pollen sterility status of 148 exotic rice germplasm was assessed at flowering stage. Sixteen genotypes showed 100% pollen sterility status which was considered as completely male sterile lines (A-line). Sixteen genotypes were also identified as completely fertile due to 80% and above pollen and spikelet fertility. For identification of proper maintainer lines, the identified 16 CMS lines chance crossed with established known maintainer lines viz. IR 58025B, IR 62829B, GAN46B, IR 68888B and BRRI1B. Based on pollen male sterility status of the F1s lines it was indicated that 10 out of 16 were maintained by IR 58025B line, 8 CMS lines were maintained by IR 62829B, three CMS were maintained by IR 68888B and one CMS line was maintained by GAN46B and BRRI1B. Restoration potentiality of identified 16 suspected restorer genotypes were assessed throgh judgement of pollen and spikelet fertility of F1s developed through crossing with five standard CMS lines. Based on 80% and above pollen and spikelet fertility of F1s, seven suspected restorers were identified as restoerer against IR 58025A, two against GAN 46A, five against IR 62829A and two against IR 68888A.

ABSTRACTIn radish, male sterility studies mainly focus on cytoplasmic male sterility (CMS) while genetic male sterility (GMS) has received less attention. To explore global gene expression profiles and identify genes associated with GMS, we performed de novo and comparative transcriptome analyses of flower buds from a GMS line and maintainer (fertile) line. Our study produced approximately 700 million paired-end reads, which were assembled into 115,284 unigenes with an average length of 1,196 bp and N50 of 1,729 bp. These unigenes were extensively annotated, with nearly 60% of them having homologs in the databases and 50% assigned with Gene Ontology (GO) terms. A total of 3,390 differentially expressed genes (DEGs) between the GMS line and the maintainer line were identified, with 2,645 down-regulated and 745 up-regulated in the GMS line. GO analyses showed biological processes related to plant reproductive development such as gametophyte development, pollen wall assembly, pollen development and pollen exine formation, as well as those related to cell wall development such as cell wall oganization and cell wall biogenesis, were significantly different in these two lines. The transcriptome assembly and comparative transcriptome analysis provides novel insights into the GMS in radishfacilitating the application of GMS in radish hybrid breeding.Abbreviation: 136S, the male sterile line in this article; 136F, the corresponding maintainer line(fertile); AGL, AGAMOUS-LIKE gene; CMS, cytoplasmic male sterility while; COG, Clusters of Orthologous Groups; DEGs, differentially expressed genes, FPKM, Fragments Per Kilobase of transcript per Million mapped reads; GMS, genetic male sterility; GO, gene ontology; KEGG, Kyoto Encyclopedia of Genes and Genomes, NPGs, non-pollen genes

Ogura cytoplasmic male sterility (CMS) lines are widely used breeding materials in cruciferous crops and play important roles in heterosis utilization; however, the sterility mechanism remains unclear. To investigate the microspore development process and gene expression changes after the introduction of orf138 and Rfo, cytological observation and transcriptome analysis were performed using a maintainer line, an Ogura CMS line, and a restorer line. Semithin sections of microspores at different developmental stages showed that the degradation of tapetal cells began at the tetrad stage in the Ogura CMS line, while it occurred at the bicellular microspore stage to the tricellular microspore stage in the maintainer and restorer lines. Therefore, early degradation of tapetal cells may be the cause of pollen abortion. Transcriptome analysis results showed that a total of 1287 DEGs had consistent expression trends in the maintainer line and restorer line, but were significantly up- or down-regulated in the Ogura CMS line, indicating that they may be closely related to pollen abortion. Functional annotation showed that the 1287 core DEGs included a large number of genes related to pollen development, oxidative phosphorylation, carbohydrate, lipid, and protein metabolism. In addition, further verification elucidated that down-regulated expression of genes related to energy metabolism led to decreased ATP content and excessive ROS accumulation in the anthers of Ogura CMS. Based on these results, we propose a transcriptome-mediated induction and regulatory network for cabbage Ogura CMS. Our research provides new insights into the mechanism of pollen abortion and fertility restoration in Ogura CMS.

Development of New InDel Marker to Detect Genotypes of Rf-1a Conferring Fertility Restoration of BT-Type Cytoplasmic Male Sterility in Rice

The genusAlternaria(Nees) is a major phytopathogen responsible for diseases. In Sesame, Alternaria alternatainfects capsules during the rainy season (August-September) in Northeast China, inducing lesion expansion and compromising seed yield and quality. To investigate the molecular response of Sesame to Alternaria alternata infection, transcriptome analyses of "Liaozhi No. 9" were conducted on the pericarp and seeds of sesame capsules before and after A. alternata infection. For the data quality control, the GC content and the proportion of Q30 bases indicated that the sequencing quality was good. The correlation analysis and principal component analysis (PCA) among sample groups demonstrated that the biological replicates had high similarity and there were obvious differences between groups. Differential gene expression analysis was performed using DESeq2. There were 5892 and 9120 differentially expressed genes in the pericarp and seeds, respectively, and among them, 2788 were the same differential genes. GO functional annotation was carried out on the differentially expressed genes, and the results showed that the differential genes were mainly enriched in biological processes (cellular processes, metabolic processes), molecular functions (catalytic activity and binding), and cellular components (cellular anatomical entities and intracellular components). KEGG metabolic pathway analysis revealed that metabolic pathways such as plant hormone signal transduction and plant-pathogen interaction were enriched after the infection of A. alternata. The genes in the jasmonic acid pathway (MYC2) and in the salicylic acid pathway (NPR1 and TGA) of the plant hormone signal transduction pathway were induced after A. alternata infection. Meanwhile, the genes in plant-pathogen interaction pathway, such as CML, CDPK and CNCGs were also induced after the infection. This study indicates that sesame capsules respond to the infection of Alternaria alternata through the genes in the plant hormone signal transduction pathway and the plant-pathogen interaction metabolic pathway, providing a theoretical basis for the subsequent research on sesame disease-resistant breeding and its molecular mechanism.

Atherigona soccata (Rondani) (Diptera: Muscidae) is one of the most important pests of sorghum, Sorghum bicolor (L.) Moench, in Asia, Africa, and the Mediterranean Europe. Exploitation of cytoplasmic male sterility (CMS) for hybrid production has resulted in considerable narrowing of the genetic base and may increase the vulnerability of this crop to insect pests. Therefore, we studied the expression of different mechanisms of resistance in sorghum to A. soccata in CMS (A) and maintainer (B) lines of 12 genotypes under field and greenhouse conditions. The CMS lines of A. soccata-resistant genotypes were preferred for oviposition (78.5 versus 71.5% plants with eggs) and suffered greater deadheart incidence (47.6 versus 41.6%) than the corresponding maintainer lines, whereas such differences were not apparent in CMS lines belonging to the susceptible genotypes (92.7 versus 92.3% plants with eggs and 75.6 versus 74.6% deadhearts) under multichoice field conditions. Similar differences also were observed under controlled conditions in the greenhouse. The larval period (9.0 versus 8.8 d) and pupal mortality (18.4 versus 13.4%) were greater on maintainer lines than that on the CMS lines in the resistant group. The male and female pupal weights, fecundity, and antibiosis index were greater on the CMS than on the maintainer lines. The maintainer lines showed better recovery resistance than the CMS lines, but no such differences were observed in tiller deadhearts. The differences in susceptibility to A. soccata were greater in the A. soccata resistant CMS and maintainer lines than in the CMS and maintainer lines belonging to susceptible genotypes. Conversion of A. soccata-resistant genotypes into alternate less susceptible cytoplasmic backgrounds may be undertaken for developing sorghum hybrids with stable resistance to A. soccata.

Expression profiles of a cytoplasmic male sterile line of Gossypium harknessii and its fertility restorer and maintainer lines revealed by RNA-Seq