Disruption of Energy Metabolism and Reactive Oxygen Species Homeostasis in Honglian Type-Cytoplasmic Male Sterility(HL-CMS)Rice Pollen

Honglian type-cytoplasmic male sterility(HL-CMS)is caused by the inter-communication between the nucleus and mitochondria.However,the mechanisms by which sterility genes regulate metabolic alterations and changes in mitochondrial morphology in the pollen of HL-CMS remain unclear.In this study,we compared the morphological differences between the pollen of the male sterile line YA and the near-isogenic line NIL-Rf6 using hematoxylin-eosin staining and 4ʹ,6-diamidino-2-phenylindole(DAPI)staining.HL-CMS is characterized by gametophytic sterility,where the aborted pollen grains are empty,and the tapetal layer remains intact.Transmission electron microscopy was employed to observe mitochondrial morphological changes at the microspore stage,revealing significant mitochondrial alterations,characterized by the formation of'large spherical mitochondria',occurred at the binucleate stage in the YA line.Additionally,metabolomics analysis revealed decreased levels of metabolites associated with the carbohydrate and flavonoid pathways.Notably,the decrease in flavonoids was found to contribute to an elevation in reactive oxygen species(ROS)levels.Therefore,we propose a model in which rice fertility is modulated by the levels of pollen carbohydrates and flavonoid metabolites,with impaired mitochondrial energy production and reduced flavonoid biosynthesis as the main causes of ROS accumulation and pollen abortion in rice.

Advances in DNA methylation and its role in cytoplasmic male sterility in higher plants

The impact of epigenetic modifications like DNA methylation on plant phenotypes has expanded the possibilities for crop development.DNA methylation plays a part in the regulation of both the chromatin structure and gene expression,and the enzyme involved,DNA methyltransferase,executes the methylation process within the plant genome.By regulating crucial biological pathways,epigenetic changes actively contribute to the creation of the phenotype.Therefore,epigenome editing may assist in overcoming some of the drawbacks of genome editing,which can have minor off-target consequences and merely facilitate the loss of a gene’s function.These drawbacks include gene knockout,which can have such off-target effects.This review provides examples of several molecular characteristics of DNA methylation,as well as some plant physiological processes that are impacted by these epigenetic changes in the plants.We also discuss how DNA alterations might be used to improve crops and meet the demands of sustainable and environmentally-friendly farming.

Morphological and cytological assessments reveal pollen degradation causes pollen abortion in cotton cytoplasmic male sterility lines

Background Understanding the mechanism of male sterility is crucial for producing hybrid seeds and developing sterile germplasm resources.However,only a few cytoplasmic male sterility(CMS)lines of cotton have been produced due to several challenges,like inadequate variation of agronomic traits,incomplete sterility,weak resilience of restorer lines,and difficulty in combining strong dominance.Therefore,the morphological and cytological identification of CMS in cotton will facilitate hybrid breeding.Results Two F_(2) segregating populations of cotton were constructed from cytoplasmic male sterile lines(HaA and 01A,maternal)and restorer lines(HaR and 26R,paternal).Genetic analysis of these populations revealed a segregation ratio of 3:1 for fertile to sterile plants.Phenotypic analysis indicated no significant differences in traits of flower bud development between sterile and fertile plants.However,sterile plants exhibited smaller floral organs,shortened filament lengths,and anther atrophy on the flowering day in comparison with the fertile plants.When performed scanning electron microscopy(SEM),the two F_(2) populations revealed morphological variations in the anther epidermis.Cellular analysis showed no significant differences in pollen development before pollen maturation.Interestingly,between the pollen maturation and flowering stages,the tapetum layer of sterile plants degenerated prematurely,resulting in abnormal pollen grains and gradual pollen degradation.Conclusion The results of this study suggest that fertility-restoring genes are controlled by a single dominant gene.Sterile plants exhibit distinctive floral morphology,which is characterized by stamen atrophy and abnormal anthers.Pollen abortion occurs between pollen maturity and flowering,indicating that premature tapetum degradation may be the primary cause of pollen abortion.Overall,our study provides a theoretical basis for utilizing CMS in hybrid breeding and in-depth investigation of the dominant configuration of cotton hybrid combinations,mechanisms of sterility,and the role of sterile and restorer genes.

ClBBM and ClPLT2 function redundantly during both male and female gametophytes development in watermelon

As a prerequisite for sexual reproduction,gametophyte development is an interesting process involving cell proliferation,differentiation and specialization.Gametogenesis has been extensively explored in model plants,but the regulatory mechanism of gametophyte development largely remains unknown in Cucurbitaceae species.In present study,we have shown that watermelon ClBBM and ClPLT2,two AP2/ERF transcription factors,participated in both male and female gametophyte development.Clbbm and Clplt2 single mutants resembled wild-type phenotypes in both vegetative and reproductive development.But Clbbm/Clplt2 double mutant showed partial pollen abortion and bore less seeds comparing to WT.Our results indicated that the abnormal pollen grains were caused by premature tapetum degeneration,and reduced seed-set was due to faulty embryo sac development.ClBBM and ClPLT2 were expressed in FG4 embryo sacs,and their transcripts were also detectable in the tapetum and microspore of stage 9 anthers,which was consistent with developmental stages of defective phenotypes observed in double mutant.The expression of genes essential for tapetum development,ClATM1,ClAMS,ClMS1 and ClMS188,was decreased in Clbbm/Clplt2 double mutants.Moreover,the transcriptome analysis indicated that ClBBM and ClPLT2 participated in tapetum and pollen wall development by regulate cell cycle,transmembrane transport,glucan and cellulose metabolic process.Collectively,ClBBM and ClPLT2 were functionally redundant in regulating gametophyte development in watermelon,and their functions differ from their homologous genes in model plant Arabidopsis.

RAPD and ISSR Markers of Fertility Restoring Gene for Aegilops kotschyi Cytoplasmic Male Sterility in Wheat

LK783 was found to be a good fertility restorer for K-type male sterility of wheat (Triticum aestivum L.). RAPD and ISSR (inter-simple sequence repeat polymorphism) markers were employed to map the major restoring gene in LK783. Maintainer and restorer DNA pools were established using the extreme sterile and fertile plants among KJ5418A//911289/LK783 F 1 population, respectively. Four hundred and eighteen RAPD primers and 33 ISSR primers were used for screening polymorphisms between the two pools, and amplification bands using a RAPD primer of OPK18 and an ISSR primer of UBC-845 were found polymorphic between the two pools. Linkage analysis showed that OPK18 450 and UBC-845 800 were linked to the restoring gene in LK783. The distance between the restoring gene and OPK18 450 was (15.07±6.28) cM (centiMorgan), with the distance between the restoring gene and UBC-845 800 being (8.20±4.85) cM. The marker of UBC-845 800 was located on chromosome 1BS by amplifying nulli-tetrasomics and 1B ditelosomics of Chinese Spring with the primer of UBC-845, indicating that the restoring gene in LK783 was located on 1BS. The breeding for new fertility restorer lines of K-type cytoplasmic male sterility of wheat would be facilitated by using the two markers.

Microtubule Structure and Male Sterility in a Gene-Cytoplasmic Male Sterile Line of Rice, Zhen Shan 97A

Histological changes that occur during microsporogenesis are documented in a gene-cytoplasmic male sterile rice ( Oryza saliva L.) line, Zhen Shan 97A, its maintainer line, Zhen Shan 97B, and the restorer line, Ce64 of a Mine hybrid rice production system. In the restorer line, Ce64, the developing microsporocytes have dense cytoplasm and a distinct set of circumferential microtubules around the nucleus. Successive cytokinesis results in the formation of tetrads. The microtubules within the cells of tetrads and microspores radiate from the surface of the nucleus towards the outer edge of the cytoplasm. Subsequent pollen development is normal. During the course of microspore formation tubulin speckles can be found in the cytoplasm. The general pattern of development and microtubule organization in the maintainer lined Zhen Shan 97B, is similar to Ce64, except that a few more tubulin speckles appear during microspore formation. In the case of the mate sterile line, Zhen Shan 97A, a number of abnormalities can be discerned during early microsporogenesis. These include vacuoles forming within the developing microsporocyte and faintly stained microtubules with no defined distribution pattern. Prominent tubulin speckles are common within the cytoplasm. For those microsporocytes that undergo meiosis, no defined organizational patterns of microtubules can be found within the tetrad. All microspores abort soon after. Abnormalities and defects in microtubule organization observed in Zhen Shan 97A showed that complex interactions between the cytoplasm and the nucleus began at very early stage of microsporocyte development.

Construction and Identification of HSP70 Antisense RNA Expression Vector for Genetic Engineering Male Sterility in Plant

[ Objective] In order to study the relation between the HSPTO gene and male sterility of plant further. [ Methods ] Anther specific expression promoter Osg6B of rice was coloned by PCR then connected with HSP70 antisense fragment to construct HSPTO antisense expression vector. The expression vector was identified by PCR experiment and enzyme digestion. [ Result] The sequence of coloned Osg6B promoter had 97% homology to the published sequence, and the cis-regulatory element in promoter area was integrated. HSP70 antisense expression vector driven by the promoter Osg6B was confired by colony PCR and enzyme digestion. [ Conclusion] The construction of expression vector would lay solid foundation for utilization of genetic engineering male sterility of plant.

Research of Fertility Standard on Male Sterility of Lycium chinensis

Botanical features of "YX-1" reproductive organ and bearing rate by number of pollen granules in pollen sacs of Lycium chinensis was observed and researched through field investigation, smear preparation and paraffin sections. YX-1 of Lycium chinensis was determined as incomplete male sterility and further classified into complete sterile type, sterile type I and sterile type II. The fertility standard of male sterility of Lycium chinensis was preliminarily proposed to establish a basis for crossbreeding of male sterility of Lycium chinensis.

Study on the Agroclimatic Division of Thermosensitive Genetic Male Sterility Rice Breeding

Using the meteorological data and geographical information during January-March（1961-2010） accumulated by 18 stations of Hainan,a suitability zoning map for thermo-sensitive genetic male sterility（TGMS） rice was made by GIS technology based on temperature indicators required by TGMS rice during fertility sensitive period and heading-flowering period,aiming to provide reasonable layout and scientific basis for sustainable development of TGMS rice in Hainan Island under the background of global warming.The results indicated that the suitable planting zones covered the south regions of Wuzhishan,Jianfengling and Diaoluoshan;subordinate suitable zones expanded northward to central parts of Wuzhishan,Dongfang,Changjiang,Qiongzhong and southern parts of Wanning;the other regions were unsuitable for breeding.

Genetic Study on Barley Male Sterility

[Objective] The aim was to research the genetic mechanism of barley male sterility, and provide the theoretical reference for breeding strong heterosis hybrid of barley. [ Method] Fertility segregation phenomenon, morphological characteristics and main agronomic characters of male sterility character of 4 kinds of barley male sterility materials 2001 - 17, 2001 - 37, 2001 - 84 and 2001 - 116 and their dedved lines were observed and researched.[ Result] Barley male sterility existed genetic phenomenon of single dominant nuclear gene caused by environmental stimulation, its sterility controlled by MS, the sterile genotype was MSms. There was stable nucleo-cytoplasmic inheritance on barley male sterility which was controlled by cytoplasmic mate sterile gene S and nuclear gene rr, and its genotype was S（rr）, this male sterility belonged to CMS type sterility. Temperature had no effect on fertility. [ Conclusion] There was stable nucleo-cytoplasmic inheritance on barley male sterility, this result played a positively promoting role in barley practical production.

Investigation on the Sterility Changeover of Male Sterility Line CMS7311 in Heading Chinese Cabbage

Sterility changeover induced by low temperature in male sterility line CMS7311 of heading Chinese cabbage ( Brassica campestris L. ssp. penkinsis) was investigated under constant temperature, day/night alternating temperature in light-growth-incubator and outdoor natural day/night alternating temperature respectively. It showed that the sterility of CMS7311 has significant change with temperature. The constant temperature of 6 - 12 degreesC was suitable to induce the changeover of sterility (CGS) of CMS7311 ; the shortest treatment time for CGS induction at this temperature was 3 d. The significant value of sterility changeover of CMS7311 occurred at the constant temperature of 6 - 9 degreesC after treatment for 6 - 9 d. The intensity of CGS was stronger under constant temperature than under alternate temperature, but the shortest treatment time inducing CGS has no difference between them. When treated for 9 d under the day/night alternating temperature of 11/7 degreesC, its sterility could also turn to normal fertility (near to sterile grade I or zero). In normal condition, the interval from the end of temperature treatment to the beginning of CGS induction was 10 - 16 d. In the condition of natural day/night alternating temperature, the lower limit of temperature for effective CGS induction was at the daily mean temperature ( DMT) of 3. 5 - 6. 8 degreesC, and the shortest treatment time which could induce the sterility changeover was also longer than that under constant temperature, being 11d at DMT of 1.5 - 3.1 degreesC. In addition, for the apical dominance, the flower bud on the first lateral branch showed the CGS 8 - 10 d later than the flower bud on the main stem under the same temperature with the same days of treatment. The intensity of CGS of CMS7311 on the first lateral branch was also weaker than the flower bud on the main stem.

Creation and Genetic Analysis of a Male Sterility Mutant in Panicum miliaceum

Heterosis plays an important role in the development of new crop varieties with high-yielding, good-quality and biotic/abiotic stresses while male sterile line de- velopment is the key step to determine the success of heterosis utilization. A male sterile mutant, M207A was created in proso millet （Panicum mi/iaceurn, 2n=4x=36） for the first time using 60Co-y ray mutagenesis. Fertility identification and genetic analysis were carried out to characterize the mutant for its possible use for hetero- sis utilization in proso millet. First the sterility was investigated using both field sur- vey and indoor pollen microscopy identification. Then Pollinated by normal fertile proso millet cultivars, F1 and F2 populations from the mutant were obtained. Mean- while primary genetic analysis was also conducted using above populations in dif- ferent experimental sites, seasons and years. The results showed that the male sterile plant exhibited closed glumes, browning and dry anthers with few normal pollens. The sterility was stable and sterility rate was above 95% on average. The segregation ratio of fertile to sterile plants was 35：1 in the fertile selfing F2 popula- tion indicating that the mutant was a genic male sterility belonging to a pollen-less type controlled by a single recessive gene. The creation of the mutant, M207A can play a key role for heterosis utilization in proso millet.

Breeding and Cultivation Technique of Recessive Genic Male Sterility Hybrid Wheat New Variety Xinan 112

Xinan 112 is the hybrid combination（11S12） by wheat recessive genic male sterility line 2011Z1 （08L5070） and restoring line K152-2, particpating in the regional test of 2011-2012, 2012-2013 and production test of 2013-2014 in Chongqing. The results showed that 3 years average yield was 4 167.5 kg/hm2, which was increased 10.5% compared with CK Yumai 7, 1 000-grain weight was 45.8 g, which was 1.7 g heavier than the control; grain number per ear was 39.3, more than 1.6 grains compared with CK. The results of 2 years quality determination were： bulk density of 811 g/L, falling number of 353 s, crude protein of 15.15%, wet gluten of 31.2%, water adsorption of 62.9 ml/100 g, formation time of 5.5 min, stable time of 6.5 min, softening degree of 90 F.U., and powder quality coefficient of 96 mm. Therefore, Xinan 112 is a high yield and high protein gluten wheat variety and is suitable for cultivated in Chongqing and climate contion similar areas.

Technical Specification for Producing Hybrid Pepper Seed Using Three-line Male Sterility

This paper described the whole process of three line hybrid pepper seed production in detail, including requirement of the seed production base, parent culti- vation, field management, and specified the key operation techniques in seed pro- duction, such as parental impurity removal to preserve pure state, pollen collection, pollination and seed collecting essentials. This specification is of guiding significance for the production of hybrid pepper seed and ensuring the purity of hybrid pepper seed.

Breeding and Application of a Japonic Rice Cytoplasmic Male Sterility Line, E-Jing A

The Chaling wild abortive type （CL-type） cytoplasmic male sterile line, E-Jing A, was characterized by stable sterility with a sterile plant percentage of 100%, a pollen sterile degree of 98% with dominated typical abortive pollens, and showed the characteristics of good outcrossing with a stigma exsertion rate of 48.2% and an outcrossing rate varied from 30% to 50%, good combining ability, and fine grain quality. E-Jing A was sensitive to temperature. Its duration from sowing to heading was 78-90 d when it was sown from mid May to early June at Wuhan. The breeding procedure of E-Jing A and its characteristics, relationship of restoring and maintaining were introduced.

Research Advances in Molecular Mechanism of Thermo-sensitive Genic Male Sterility in Plants

Thermo-sensitive genic male sterile （TGMS） lines have specific superiority in heterosis utilization of crops. So far, thermo-sensitive genic male sterile lines have been found in many plants and are widely used in two-line hybrid breeding. With the rapid development of molecular biology, the molecular nature of thermo-sensitive genic male sterility has been revealed, which lays the foundation for further devel- opment and utilization of thermo-sensitive genic male sterile lines. In this study, the molecular mechanisms of fertility conversion of plant thermo-sensitive genic male sterile lines were reviewed from gene molecular mapping and gene differential ex- pression, and the mechanisms of gene differential expression in thermo-sensitive genic male sterile lines were further discussed.

Mapping of Fertility Restoring Gene for Aegilops kotschyi Cytoplasmic Male Sterility in Wheat Using SSR Markers

LK783 was found to be a good fertility restorer for K-type male sterility of wheat. Microsatel-lite markers were employed to map the major restoring gene in LK783. Maintainer and restorer DNA pools were established using the extreme sterile and fertile plants among (KJ5418A//911289/LK783)F1 population, respectively. Seventy-nine sets of SSR primers were screened for polymorphism between the two pools, 6 of which were found polymorphic. Linkage analysis showed that Xgwm11, Xgwm18, Xgwm264a and Xgwm273 were linked to the restoring gene in LK783, while Xgwm11, Xgwm18 and Xgwm273 were co-segregated. The distance between the Rf gene in LK783 and the three co-segregated markers was 6.54 ± 4.37 cM, the distance between Rf gene and Xgwm264a was 5. 71 ± 4.10 cM. The four SSR markers were located on chromosome IBS by amplifying the DNA of nulli-tetrasomics and ditelosomics of CS with the 4 sets of primers, indicating that the major restoring gene in LK783 was located on IBS, but the relative location of the gene was different from Rfv1, allelism of the two genes should be further investigated. The breeding for new fertility restorer lines of K-type cytoplasmic male sterility in wheat would be facilitated by using the four polymorphic markers.

Genetic Analysis and Primary Mapping of pms4, a Photoperiod-Sensitive Genic Male Sterility Gene in Rice (Oryza sativa)

To understand the genetic characteristics of a new photoperiod-sensitive genic male sterile line Mian 9S, some reciprocal crosses were made between Mian 9S and six indica rice materials, Yangdao 6, Luhui 602, Shuihui 527, Mianhui 725, Fuhui 838 and Yixiang 1B. Genetic analysis results suggested that the photoperiod-sensitive genic male sterility （PGMS） of Mian 9S was controlled by a single recessive nuclear gene. Thus, the F2 population derived from the cross of Yangdao 6/Mian 9S was used to map the PGMS gene in Mian 9S. By using SSR markers, the PGMS gene of Mian 9S was mapped on one side of the markers, RM6659 and RM1305, on rice chromosome 4, with the genetic distances of 3.0 cM and 3.5 cM, respectively. The gene was a novel PGMS gene and designated tentatively as pms4. In addition, the application of the pms4 gene was discussed.

Mitochondrial localization of ORF346 causes pollen abortion in alloplasmic male sterility

The Nsa cytoplasmic male sterility(CMS) system confers stable male sterility and offers great potential for production of hybrid seeds in oilseed rape. However, genes responsible for male sterility in Nsa CMS have not been identified. By mitochondrial genome sequencing of Nsa CMS and its maintainer line,we identified in an Nsa CMS line several chimeric genes encoding hypothetical proteins harboring transmembrane domains. One novel chimeric gene orf346 showed high identity with cox1 at the 50 terminal region and was co-transcribed with nad3 and rps12 genes. Transgenic plants of orf346 fused with or without mitochondrial targeting peptide conferred complete male sterility in Arabidopsis. ORF346 was mitochondrion-localized. Expression of orf346 in Escherichia coli inhibited bacterial growth, with excessive accumulation of reactive oxygen species and decreased ATP content. These results reveal a link between the newly identified mitochondrial gene orf346 and the abortion of Nsa CMS. Inadequate energy supply and excessive accumulation of reactive oxygen species may account for pollen abortion in Nsa CMS plants.

Identification of a locus associated with genic male sterility in maize via EMS mutagenesis and bulked-segregant RNA-seq

Genic male sterility(GMS) is one of the most important resources for exploiting heterosis in crop breeding, so that identifying genomic loci regulating GMS is desirable. However, many regulatory genes controlling GMS have not yet been characterized in maize, owing partly to a lack of genetic materials. We generated a recessive male-sterile maize mutant in the Jing 724 genetic background via ethyl methanesulfonate treatment, and found the male sterility to be due to a single gene mutation. Bulk-segregant RNA sequencing of three replicates indicated that one genomic region located at the end of chromosome 4 was associated with the observed mutant phenotype. Among genes with nonsynonymous mutations,Zm00001 d053895(bHLH51) showed abolished expression in the sterile bulks and was annotated as a bHLH transcription factor orthologous to Arabidopsis AMS, suggesting an association with the male sterility of the mutant. Kompetitive Allele-Specific PCR assays further validated the exclusive correlation of male sterility with the single C-to-T mutation in the fifth exon. The new maize mutant and the potential SNP locus provide novel genetic material for investigating the molecular mechanism underlying tapetal development and may facilitate the improvement of hybrid production systems.