Cytoplasmic Sterility Research Articles

Insights into cellular crosstalk regulating cytoplasmic male sterility and fertility restoration.

Cytoplasmic male sterility has been a popular genetic tool in development of hybrids. The molecular mechanism behind maternal sterility varies from crop to crop. An understanding of underlying mechanism can help in development of new functional CMS gene in crops which lack effective and stable CMS systems. In crops where seed or fruit is the commercial product, fertility must be recovered in F1 hybrids so that higher yield gains can be realized. This necessitates the presence of fertility restorer gene (Rf) in nucleus of male parent to overcome the effect of sterile cytoplasm. Fertility restoring genes have been identified in crops like wheat, maize, sunflower, rice, pepper, sugar beet, pigeon pea etc. But in crops like eggplant, bell pepper, barley etc. unstable fertility restorers hamper the use of Cytoplasmic genic male sterility (CGMS) system. Stability of CGMS system is influenced by environment, genetic background or interaction of these factors. This review thus aims to understand the genetic mechanisms controlling mitochondrial-nuclear interactions required to design strong and stable restorers without any pleiotropic effects in F1 hybrids.

Reconsidering the Lessons Learned from the 1970 Southern Corn Leaf Blight Epidemic.

The southern corn leaf blight epidemic of 1970 caused estimated losses of about 16% for the U.S. corn crop, equivalent to about $8 billion in current terms. The epidemic was caused by the prevalence of Texas male sterile cytoplasm (cms-T), used to produce most of the hybrid corn seed planted that year, combined with the emergence of a novel race of the fungus Cochliobolus heterostrophus that was exquisitely virulent on cms-T corn. Remarkably, the epidemic lasted just a single year. This episode has often been portrayed in the literature and textbooks over the last 50 years as a catastrophic mistake perpetrated by corn breeders and seed companies of the time who did not understand or account for the dangers of crop genetic uniformity. In this perspective article, we aim to present an alternative interpretation of these events. First, we contend that, rather than being caused by a grievous error on the part of the corn breeding and seed industry, this epidemic was a particularly unfortunate, unusual, and unlucky consequence of a technological advancement intended to improve the efficiency of corn seed production for America's farmers. Second, we tell the story of the resolution of the epidemic as an example of timely, meticulously applied research in the public sector for the public good.

Analysis of the cytohistological phenotype of anthers in sunflower with ASI-AG3 in the context of the PET1 mytotype and the allelic genotype for the Rf1 gene

Microsporogenesis and male gametogenesis are processes that, although under tight morphogenetic control, can often be affected by deviations of internal or external stimuli causing androsterility. The article examines the cytohistological phenotypes of sunflower anthers with gibberellin induced androsterility in plants with different mitotype and allelic genotype for the Rf1 gene, in comparison with the cytohistological phenotypes of fertile and sterile anthers (ASC-PET1). The treatment with gibberellic acid (AG3, 0.01%) was carried out by spraying the inflorescence at the budding stage. Cytological samples and those with semi-thin anther sections were analysed by light microscopy. It was determined that the gametocidal effects induced by AG3 associated with degenerative and early apoptotic events in tapetum cells were more severe in plants with sterile cytoplasm and nuclear Rf1 genes compared to those with fertile cytoplasm and no fertility-restoring genes. In plants with ASC, the exogenous gibberellin stimulus does not cause any visible compensatory events of the sterile phenotype, on the contrary, the effects of microsporogenesis inhibition are more temporally advanced, with sporogenous and sporophytic tissue cells showing various lesions in the premeiosis. The general common and differentiation characteristics found at the androsteril phenotypes suggest interactions between orfH522 and Rf1 gene expression products of negative feedback type in untreated plants and positive feedback type in AG3-treated ones. The research is relevant in the context of elucidating the physiological regulatory processes of the sensitivity of male gametophyte development to various stress conditions, which prospectively makes possible regulatory intervention in plants with different androsterility/androfertility control systems.

Cyto-swapping in maize by haploid induction with a cenh3 mutant.

Maize mutants of the centromeric histone H3 (CENP-A/CENH3) gene can form haploids that inherit only chromosomes of the pollinating parent but the cytoplasm from the female parent. We developed CENH3 haploid inducers carrying a dominant anthocyanin colour marker for efficient haploid identification and harbouring cytoplasmic male sterile cytoplasm, a type of cytoplasm that results in male sterility useful for efficient hybrid seed production. The resulting cytoplasmic male sterility cyto-swapping method provides a faster and cheaper way to convert commercial lines to cytoplasmic male sterile compared to conventional trait introgression.

Commercial suitability and characterization of newly developed Erucastrum canariense (Can) sterile cytoplasm based cytoplasmic male sterile (CMS) lines in Indian cauliflower

The study presents the first to characterize novel Erucastrum canarianse Webb and Berthel (or Can) sterile cytoplasm-based CMS lines in Indian cauliflower (Brassica oleracea var. botrytis L.) and investigating their commercial suitability. Eleven Can-based CMS lines were examined for 12 agro-morphological and yield traits,18 floral traits, four seed yield traits together with three each of the Ogura (source: wild Japanese Radish) and Tour (Source: Brassica tournefortii) cytoplasms. All of the recorded floral and seed traits showed significant (P > 0.05) differences between the CMS lines of each group. Agro-morphological and yield traits in CMS lines and their maintainers, however, were non-significantly different. All the Can- and Ogura-based CMS lines showed flowering and appropriate seed formation by natural cross-pollination. Only two Tour cytoplasm-based CMS lines, Tour (DC-41-5) and Tour (DC-67), produced the smallest malformed flowers and stigma. The highest seed yield per plant in CMS lines was in Ogu (DC-98-4) and the lowest in Tour (DC-67). P14 and P15, two polymorphic mtDNA markers, were discovered for the Can CMS system for early detection. Five primers (ITS5a-ITS4, atpF-atpH, P16, rbeL and trnL), along with their maintainers, were sequenced and aligned to detect nucleotide changes including as additions and or deletions at different positions. The newly introduced E. canariense sterile cytoplasm-based CMS system in cauliflower is the subject of the first comprehensive report, which emphasises their potential as a further stable and reliable genetic mechanism for hybrid breeding.

Development and Investigation of HRM Markers to Discriminate Two Ogura Cytoplasmic Male Sterility Restorer Genes in Radish

Ogura male sterile cytoplasm is widely used for radish breeding. In this study, high-resolution melting (HRM) markers associated with Rft and Rfo, major restorer-of-fertility genes in Ogura cytoplasmic male sterility (CMS) in radish, were developed. Genetic mapping was carried out using F2 populations derived from crosses between male-sterile Ogura CMS lines and male-fertile lines. Identification of the Rft and Rfo loci was achieved through SNP-based genotyping and linkage grouping. HRM markers were subsequently developed based on flanking sequences of SNPs linked to these loci. For the Rft gene, a set of 117 SNPs was selected within a candidate region on chromosome 5, and 14 HRM markers were successfully developed. Genotyping of F2 showed high correlation between three markers and the phenotype. Regarding the Rfo gene, a set of 27 HRM markers was designed based on flanking sequences of SNPs located on chromosomes 9 and 0. Genotyping in the Rfo segregating population identified a single marker, RSRF27, that accurately distinguished the male sterility phenotype. Validation of the developed markers was performed in populations containing both Rft and Rfo genes, confirming their utility for genotyping and demonstrating that these two genes independently contribute to male sterility recovery. Overall, this study provides HRM markers that can be used for genotyping Rft and Rfo and contributes to a deeper understanding of male sterility restoration mechanisms in Ogura CMS.

Correlation between the response of maize lines to CMS and their origin

Transferring maize hybrids to a sterile basis is the most important condition for their introduction into production. In this regard, it is important to study the behavior of the initial forms of hybrids, namely self-pollinated lines, in the sterile cytoplasm. The purpose of the current study was to estimate the response of new self-pollinated maize lines, depending on their origin, to the “M” and “C” types of CMS, to identify natural restoring agents and complete sterility fixatives. The study was carried out at the FSBSI “ARC “Donskoy” in 2021–2023. The subjects of the study were 20 new self-pollinated maize lines of various origins, 6 sources of sterility, 120 testcross maize hybrids. There was found that the response of maize lines depended on their origin. Lines belonging to the same group (by origin) usually had the same reaction to the Moldovan (“M”) and Bolivian (“C”) types of cytoplasmic male sterility (CMS). The closer the lines are related, the smaller the differences. The new self-pollinated lines of the first group ‘SL 171’, ‘SL 172’, ‘SL 174’, ‘SL 175’, ‘SL 176’ and ‘SL 177’ were characterized by complete consolidation of the type “M” of CMS (class 0). Lines of subgroup Ia were fixed, but subgroups Ib did not completely restore the type “C” of sterility. In the second group there were identified the complete ‘SL 181’, ‘SL 182’ (class 4, 5) and incomplete ‘SL 183’, ‘SL 184’ (class 2–5) reducing agents of the type “M” of sterility. There was determined that all lines of the second group perpetuated sterility of the Bolivian type (class 0). There have been identified the lines of the third group ‘SL 191’, ‘SL 192’, ‘SL 193’, ‘SL 194’, as full reducing agents of the type “M” of CMS, two of them ‘SL 191’ and ‘SL 193’ are also complete reducing agents of the type “C” of CMS (class 4, 5). All lines of the fourth group could be classified as complete fixatives (class 0, 1) of the studied types of CMS. The results obtained will improve the efficiency of work on transferring maize hybrids to a sterile basis.

Allelic haplotype combinations at the MS-P1 region, including P-class pentatricopeptide repeat family genes, influence wide phenotypic variation in pollen grain number through a cytoplasmic male sterility model in citrus.

In citrus breeding programs, male sterility is an important trait for developing seedless varieties. Sterility associated with the male sterile cytoplasm of Kishu mandarin (Kishu-cytoplasm) has been proposed to fit the cytoplasmic male sterility (CMS) model. However, it remains undetermined whether CMS in citrus is controlled by interactions between sterile cytoplasm and nuclear restorer-of-fertility (Rf) genes. Accordingly, mechanisms underlying the control of the wide phenotypic variation in pollen number for breeding germplasm should be elucidated. This study aimed to identify complete linkage DNA markers responsible for male sterility at the MS-P1 region based on fine mapping. Two P-class pentatricopeptide repeat (PPR) family genes were identified as candidates for Rf based on predicted mitochondrial localization and higher expression in a male fertile variety/selected strain than in a male sterile variety. Eleven haplotypes (HT1-HT11) at the MS-P1 region were defined based on genotyping of DNA markers. Association analysis of diplotypes at the MS-P1 region and the number of pollen grains per anther (NPG) in breeding germplasms harboring Kishu-cytoplasm revealed that the diplotypes in this region influenced NPG. Among these haplotypes, HT1 is a non-functional restorer-of-fertility (rf) haplotype; HT2, a less-functional Rf; HT3-HT5 are semi-functional Rfs; and HT6 and HT7 are functional Rfs. However, the rare haplotypes HT8-HT11 could not be characterized. Therefore, P-class PPR family genes in the MS-P1 region may constitute the nuclear Rf genes within the CMS model, and a combination of the seven haplotypes could contribute to phenotypic variation in the NPG of breeding germplasms. These findings reveal the genomic mechanisms of CMS in citrus and will contribute to seedless citrus breeding programs by selecting candidate seedless seedlings using the DNA markers at the MS-P1 region.

Assembly and Analysis of Plastomes for 15 Potato Cultivars Grown in Russia

Chloroplasts are important organelles in a plant cell, having their own DNA (cpDNA), transmitted only through the female line, and performing the function of photosynthesis. The determination of chloroplast DNA is of interest in the study of the genetic diversity and phylogeny of potatoes, and of cytoplasmic sterility, as well as for applications in biotechnology and genetic engineering. Here, we reconstructed the complete plastomes of 15 S. tuberosum potato cultivars grown in Russia. Our analysis allowed us to determine the composition and location of genes for these plastid DNAs. It was shown that the plastid genome contains both highly and low-variable regions. The region at position 63,001–68,000 nt has the highest variability. We determined the types of cpDNA based on in silico approaches: 10 cultivars have cpDNA of the W-type and 5 cultivars have cpDNA of the T-type. The genetic diversity of the plastid DNA for these potato cultivars was analyzed alongside the previously reconstructed plastomes of South American accessions, European/North American commercial cultivars and potato cultivars bred in the Ural region. The results show that plastid DNAs of the same type form clusters by sequence similarity, in agreement with previous studies.

A study on the adaptive potential of new sterile cytoplasm of Beta patula and Beta maritima L. from wild beets of the genus Beta L.: cold resistance and traits of early sugar accumulation in introduced alloplasmic lines

Purpose. Assessment of new sugar beet breeding genotypes by a set of economically valuable traits to determine their suitability for the efficiency of the process of managing the production of bioethanol with the study of the activity of photosynthesis according to the content of chlorophylls a and b, early accumulation of carbohydrates, tolerance to low temperatures, germination of apozygous seeds and using new plasma of the substituted lines of sugar beet with introduced sterile cytoplasms from wild species Beta patula and B. maritima L. and apozygous lines A9 with S vulgaris Owen's cytoplasm. Methods. Field methods were used (pollen-free regime for the reproduction of apozygotic seeds in conditions of isolation and seed monogermity of the seed plants), laboratory methods (selection of the best experimental genotypes for cold resistance, analysis of productivity and dynamics of carbohydrate accumulation). Results. Self-fertile pollen-sterile lines A9 Beta vulgaris Sxxzz rr with an apomictic method of seed reproduction of seeds (Yaltushkiv EBS) and commercial hybrids with early carbohydrate accumulation were selected for the recessive colour of the hypocotyl r-r- and gametophytic reduced parthenogenesis for hybridization according to the scheme: A9 Beta vulgaris Sxxzz rr × Beta vulgaris Mm Rr. The influence of cytoplasmic inheritance of B. maritima and B. patula on cold resistance indicators was studied and seed samples of the experimental genotypes No. 17225, 17221, 17222, 17220, 17226, 17223 were selected among substituted lines with seed germination rates at +4°C varying from 14.3 to 40.3%. Experimental genotypes of the substituted lines, grown in the conditions of pots were selected and examined by the indicators of early accumulation of sugars, which had values from 16.2 to 17.6% during four months of vegetation; photosynthetic activity with indicators of the content of chlorophyll varying from 2.06 to 1.32%, and chlorophyll b from 1.02 to 0.65%. Conclusions. Sugar beet hybrids, created with the use of cold-resistant substituted pollen-sterile lines of sugar beet, thanks to the natural evolution of the introduced cytoplasmic genome, ensure the extension of the growing season, high productivity, early accumulation of sugars and enrichment of the crop adaptation potential. The article contains the results of the study of the early carbohydrate accumulation, photosynthetic activity of selected experimental genotypes of the substituted sugar beet lines with high seed quality and 100% monogermity. Root crops of apomictic lines were selected based on R+r hypocotyl colour, generative reduced parthenogenesis, and studied for their specifics of seed set under the pollen-free regime.

Assessment of combinational ability of inbred lines of winter rye by the topcross method

Dentification perspective on combinational ability of the inbred lines of winter rye is an important stage in hybrid breeding. The purpose of researches - to estimate the general and specific combinational ability of the inbred lines of winter rye by a method of full top crossings. As a maternal form 30 homozygous lines of rye bearing sterile cytoplasm Pampas type served. As testers 4 male fertile homozygous lines used : mfН-1432, mfН-1423, mfН-1147 and mfН-1842. Test crossings carried out in 2019 on spatially the isolated plots. In total 120 simple interlinear hybrids of F1 were received. Their test carried out in 2020 on plots 8,0 m2 in 2 repetitions at norm of seeding of 500 grains on 1 m2. As the standard used population variety Valdai. Productivity of hybrids F1 varied from 5,08 to 7,55 t/hectare at average productivity in experience 6,27t/hectare. Hybrids with participation of the line msH-1185-1 were the most yielding. Their productivity varied from 6,92 to 7,48 t/hectare depending on a tester. Frequency of hybrids of F1 with authentically high competitive heterosis made 20 of 120 or 16,7 %. In dispersion of a sign of productivity the essential contribution was made both the general (GCA), and specific (SCA) combinational ability. However fell to the share of effects GCA 91,2 % of all dispersion of productivity, and to SCA share - 6,9 %. The high relation variances by GCA/SCA is pointed to rather high genetic divergence by the inbred lines taken for crossing. Authentically high GCA showed 7 sterile lines: ms H-1185-1, msH-1185-2, msН-1179, msН-1238, msН-700, msН-48, msН-1247. Among lines testers of the best on GCA there was mfН-1147. High effects of SCA showed the msН-451, msН-577, msН-842, msH-1185-2 and msН-1190 lines, and also testers of mfН-1432 and mfН-1423. It is characteristic that average productivity of hybrids with participation of lines with high GCA was authentically above, than average productivity of hybrids with participation of lines from high SCA (6,76 t/hectare against 6,24 t/hectare, tfact=3,38> t05=2,22). Significantly low GCA possessed 9 sterile lines. Positive correlation between effects of GCA lines and level of their own productivity is revealed (r=0,59, t fact =2,30> t 05=2,22)

ВЛИЯНИЕ ТИПА СТЕРИЛЬНОЙ ЦИТОПЛАЗМЫ НА СЕЛЕКЦИОННО-ЦЕННЫЕ ПРИЗНАКИ ГИБРИДОВ F1 СОРГО В РАЗЛИЧНЫХ ПО ВЛАГООБЕСПЕЧЕННОСТИ УСЛОВИЯХ

Some researchers have found the influence of sterile cytoplasm on the manifestation of biological and valuable breeding traits in sorghum. In addition, some authors detect the influence of sterile cytoplasm, while others describe the absence of differences between F1 hybrids obtained from CMS lines with the same nuclear genome and differing only in the type of sterile cytoplasm. In this context, the aim of the research was to determine the effect of sterile cytoplasms A1, A2, A3, A4, A5, A6 and meteorological conditions of growing F1 hybrids of grain sorghum on the main valuable breeding traits. In this work, F1 hybrids were obtained on the basis of CMS lines with the Karlik 4v genome and six types of sterile cytoplasm, and the Vostorg line was used as a pollinator. The studies were carried out in 2016–2018, with different hydrothermal regimes of plant growing seasons (GTK=0.51–1.01). As a result of the experiment, for the first time an increase in plant height during maturation was found in the A5 Karlik 4v/Vostorg hybrid (123.3 cm) compared to hybrids on cytoplasms A1, A2, A3, A4, A6 (118.0 cm); a decrease in the flag leaf area was found in the A3 Karlik 4v/ hybrid (104.4 cm2) compared to hybrids on cytoplasms A2, A4, A5 and A6 (130.3–136.3 cm2). On average, during the test period, hybrids on A1 and A5 cytoplasms produced a higher biomass yield (18.53–18.57 t/ha) than hybrids on A4 and A6 cytoplasms (13.76–15.91 t/ha), but the differences were not significant. At the same time, the contribution of the factor “CMS type” to the totalvariability of breeding traits ranged from 1.4 to 14.0%; “meteorological conditions” of the year – 24.0–58.9%. When breeding sorghum hybrids using genetically different types of sterile cytoplasm, it is advisable to include a CMS line on the A5 cytoplasm in the crossing according to the complex of breeding characteristics.

Methods of creating substituted lines for hybrid selection of sugar beet: plants of the species Beta vulgaris L. with new sterile cytoplasm of Beta patula L.

Purpose. Due to changing climatic conditions and the spread of new viral diseases, an important issue in beet growing is the expansion of plasma ulture using new sterile cytoplasm from wild species of the genus Beta L.
 as an alternative to Owen’s S-cytoplasm. The goal of this work was to obtain new breeding genotypes based on new cytoplasm. Methods. Cytological, biotechnological. field and laboratory. Results. Availability of recombination in the locus of grafted genes of one-year and two-year development cycles B+b- and hypocotyl colour R+r- was determined using the method of analytical crosses. Earlier we have proved the positive effect of sterility maintainer Beta vulgaris Nxxzz in replacement of the sterility maintainer genome based on our results on the selection of hypocotyl of green color r-rand two-year development cycle b-b- in the interspecific hybrids F2 obtained under the hybridization scheme Beta maritima L: ♀B1C maritima / ♂ Beta vulgaris Nxxzz (Роїк М. В., 2013). In the case of interspecific hybrids, in the conditions of sterile cytoplasm Beta patula L., only backcross offspring of the third cycle of saturating crosses were identified by one- and two-year developmental cycles. Seed bearing plants of BC3S patula of a two-year development cycle were characterized by spreading stems, multi-seeded fruits (3, 4 seeds) and sterile anthers. At the same time, monogermity and a two-year development cycle were restored against the background of sterile cytoplasm of Beta patula L. only among the seed bearing plants of the fourth cycle of saturating crosses. Recombinant genotypes with a red hypocotyl colour and a two-year development cycle BC4S patula R+r- were characterized by multigermity, chimericalness by sterility of CMS‑1 and CMS‑0 type and single- and multi-stem morphological types of anthers. The expression of CMS nuclear genes in the newly replaced BC5S patula lines was marked by high sterility rates of the CMS‑0 type; however, the fertility rates required additional rejection of multigerm seed plants. This is an indicator of the complex structure of the fertility locus in wild Beta patula L. and the special response of the nuclear genome of the sterility maintainers to the new cytoplasmic background. Experimental triploid hybrids with the new introductory sterile cytoplasm Beta patula L. were determined by high sugar content and dry matter content up to 29%. Conclusions. Alloplasmic lines based on the cytoplasm of the wild species Beta patula L are created. Their sugar content at the level of experimental triploid hybrids was 19.5% and 20.5% with a dry matter content of 25% —29%. Alloplasmic lines with a new BCC Beta patula L. recommended for use in the selection process of sugar beets.

Seed productivity of alloplasmic lines of Beta patula and B. maritima with sterile cytoplasm under the conditions of apozygotic reproduction

Purpose. Revealing the effect of the cytoplasmic genome of replaced lines with the plasma of wild species Beta patula and B. maritima L., and apozygotic lines A4–A8 with the cytoplasm of S vulgaris Owen on the main factors of apozygotic reproduction, seed productivity, germination, monogermity and sterility as affected by the genetic origin of breeding material.
 Methods. The research was carried out in the Cytogenetics Laboratory (IBCSB), the Laboratory for Apomixis and Polyploidy of the Yaltushkiv EBS, and the Laboratory for Adaptive Breeding (Veselyi Podil EBS). Apozygotic seeds were obtained under a pollen-free regime according to the IBCSB’s Methods for Spatial Isolation and Parchment Insulators. Each seed bearer plant phenotype was determined during the flowering period by pollen sterility and partial flowering. Classification of plants was performed according to Owen (1945), with identifying plants of CMS-0, Type CMS-1, and CMS-2 types. The Monogermity of seed plants was assessed visually by the presence of separate fruits on the central shoots. In 2021, the roots of the replaced lines of the Veselyi Podil EBS were planted under the conditions of a pollen-free regime in the experimental field of the IBCSB. Seed production under apozygositic conditions was studied, taking into account the number of set fruits per 10-cm segment with 5 replications for each seed bearer. Germination was determined on the 10th day and germination vigour on the 5th day.
 Results. New sources of cytoplasmic male sterility (CMS) were obtained in the cytogenetics laboratory based on a genetic model of crossbreeding analysis, using differentiation and tools according to the marker-linked genes of hypocotyl color R+r-, and one / two-year development cycle B+b-. The analyzers for the nature of sugar beet sterility were sterility maintainers, dominant homozygotes for recessive genes of anthocyanin color, development cycle, partial fertility and sterility (NBeta vulgaris Sxxzz rr bb). Monogerm pollen-sterile lines with an apomictic way of seed reproduction (Yaltushkiv EBS) (A4–A8 Beta vulgaris Sxxzz rr), selected for the dominant color of the hypocotyl R+r-, stabilized for the trait of monogermity, 100% sterility and 2x gene ploidy, were characterized by low seed productivity. High rates of apozygotic seed development (80 to 96.4% of the number of set flowers) were observed against the background of the sterile cytoplasm of Beta maritima (Turkey). In the CMS BC4S patula line, the number of set apozygotic seeds ranges from 34 ± 0.3 to 39 ± 0.42, and the indicators of degenerated flowers under apozygotic regime varied from 31.2 to 54.3%. Isolated were seed plants with high self-reproduction, such as BC4S maritima Turkey, k.2/1, k.6/2, k.6/3 and k.3/4, k.9/4 on the background of new plasma of wild species of B. patula. The phenomenon of high self-reproduction of seeds (up to 98.5−96.4% of set flowers) was detected in line 21-011 CHS BC5S patula, which was comparable with hybrids and sterility maintainers (97.5−93.1%), which is determined by a special interaction of the beet nuclear genome of sugar beet and new plasma of wild species of the genus Beta L.
 Conclusions. The apomictic way of seed reproduction ensures the shortening of the breeding scheme for sugar beet due to high seed reproduction of mother parent in substituted lines with new plasma and differentiation by gametophytic reduction of parthenogenesis using morphological marker traits.

The response of maize lines to the Paraguay type of CMS

Abstract. The development of cytosterile maize hybrids in Russia is a necessary condition for their wide implementation into production. To develop such hybrids, it’s greatly relevant to be aware how the lines react to sterile cytoplasm. The study was carried out at the Agricultural Research Center “Donskoy” (“ARC “Donskoy”) in 2010–2021. The purpose of the current study was to classify the new self-pollinated maize lines according to the composition of the fertility-restoring genes of the Paraguay (C) type of CMS, to optimize the number of analyzing test-crosses. Methods. As initial material there have been used 45 new self-pollinated maize lines and 8 sources of sterility with different genetic structure. The method of complete top-crosses there have been identified 360 maize hybrids, used for estimation of the new lines’ reaction. Results. According to the study results, the sterility-fixing lines KV 204, SP 286, DS 255, SP 207, DS 180, which had no fertility-restoring genes in the dominant state, belonged to the I class. As the natural complete constant fertility-restorers there has been recommended to use the lines of the VIII class KV 498, KV 272, KV 7/07, SP 357, RD 261, DS 295, SP 210, SP 197, DS 177, DS 188, having all three dominant genes Rf4, Rf5, Rf6 in the genotype. The incomplete sterility-fixing lines included the lines of the II–IV classes (KV 3, RD 245, SP 198, etc.). The incomplete fertility-restoring lines were the lines of the V–VII classes (KV 469, RD 331, KV 276, etc.). There has been found out that the most common lines were the lines of the V (24.4%) and VIII (22.3 %) classes. The scientific novelty of the study was an optimal number and genetic structure of the analyzers WF 9c of the V class, Lc of the VI class and W 401c of the VII class, which are necessary for crossings and allow identifying lines according to the fertility-restoring genes.

Integrated analysis of metabolome and transcriptome reveals the cytoplasmic effects of CMS-D2 on pollen fertility resulting from disrupted lipid metabolism.

Using cytoplasmic male sterility of Gossypium harknesii (CMS-D2) is an economical and effective method to produce cotton hybrids. However, the detrimental cytoplasmic effects of CMS-D2 on pollen fertility and fiber yields greatly limit the further development of three-line hybrid cotton in China. In this study, an integrated non-targeted metabolomics and transcriptome analysis was performed on mature pollens of maintainer line NB, isonuclear alloplasmic near-isogenic restorer lines NH and SH under two environments. A total of 820 metabolites were obtained, of which lipids and lipid-like molecules were the most, followed by organic acids derivatives, phenylpropanoids, and polyketides. Transcriptome analysis revealed significantly more differentially expressed genes (DEGs) in SH versus NH both in Anyang and Jiujiang, and most of the DEGs were significantly upregulated. Further KEGG analysis showed that most DEGs were enriched in the biosynthesis of unsaturated fatty acids, phenylalanine metabolism, and phagosome. Based on the weighted gene co-expression network analysis, totally 74 hub genes were also identified, of which three transcription factors, i.e., WRKY22, WRKY53, and ARF18 were significantly upregulated in SH and may play a negative regulatory role in pollen development by directly or indirectly regulating the jasmonic acid synthesis and signal transduction. Moreover, we found that the negative effects of CMS-D2 cytoplasm on pollen fertility were mainly due to disturbed lipid metabolism, especially the metabolic balance of unsaturated fatty acids, ultimately resulting in the decline of pollen fertility. Meanwhile, in the presence of CMS-D2 sterile cytoplasm, the cytoplasmic-nucleus interaction effects generated a substantial quantity of flavonoids involved in the fertility restoration process. This study preliminarily clarified some of the reasons for the negative effects of CMS-D2 cytoplasm on pollen fertility, and our results will provide an important theoretical reference for further breeding and improvement of three-line hybrid cotton in the future.

Characterization, inheritance of male sterility and development of male sterile and maintainer lines in ridge gourd (Luffa acutangula (Roxb.) L.)

Two male sterile mutants IIHRRG-12MS (long fruited) and IIHRRG-28MS (medium long fruited) were identified from the ridge gourd germplasm IIHR-12 and IIHR-28 respectively at ICAR-IIHR, Bengaluru. These two male-sterile (ms) sources were characterized by the production of rudimentary male flowers in the racemes in contrast to the bright yellow flowers with fertile pollen and healthy anthers in male fertile, monoecious plants. Using these ms lines the inheritance of male sterility was worked out, which is cytoplasmic genic male sterility (CGMS) type, with single dominant gene either in homozygous or heterozygous condition restoring male fertility in the presence of sterile cytoplasm. In order to develop F1 hybrids using male sterility, several male sterile and maintainer lines were developed in different genetic back grounds such as green/dark green fruit colour and short/medium long/long fruit length.

Transcriptomic and iTRAQ-Based Quantitative Proteomic Analyses of inap CMS in Brassica napus L.

Brassica napus inap cytoplasmic male sterility (CMS) is a novel sterile line with potential application in rapeseed hybrid breeding. Sterile cytoplasm was obtained from Isatis indigotica through somatic fusion and then recurrent backcrossing with B. napus. Previous studies have shown that inap CMS abortion occurred before the stamen primordia (stage 4–5), but the genetic mechanism of sterility needs to be studied. RNA-seq analyses were performed on the floral buds at two stages (0–5 and 6–8), before and after the formation of stamen primordium. As a result, a total of 1769 and 594 differentially expressed genes (DEGs) were detected in the CMS line compared to its maintainer line at the two stages, respectively. In accordance with the CMS phenotype, the up- and downstream regulators of the stamen identity genes AP3 and PI were up- and downregulated in the CMS line, respectively. Furthermore, isobaric tags for relative and absolute quantitation (iTRAQ) analysis showed that a total of 760 differentially abundant proteins (DAPs) were identified in flower buds at stages 0–8, and most of the proteins related to the anther development, oxidative phosphorylation, and programmed cell death (PCD) were downregulated in inap CMS. In combined transcriptomic and proteomic analysis, a total of 32 DEGs/DAPs were identified, of which 7 common DEGs/DAPs had the same expression trend at stage 0–8 of flower development. The downregulation of genes related to the energy deficiency, hormone signal transduction, and the maintenance of mitochondrial metabolic homeostasis at stage 0–5 might disturb the normal differentiation of stamen primordium, resulting in carpelloid stamen of inap CMS. The study will help provide insights into the molecular mechanism of this new male sterility.

Agro-morphological and molecular diversity analysis of new cytoplasmic male sterile lines in Indian cauliflower for their use in hybrid breeding

Cauliflower is a typical thermo-sensitive crop, highly responsive to heterosis for yield and stress tolerance. A diverse set comprising of 24 cytoplasmic male sterile (CMS) lines, developed across different maturity groups of Indian cauliflower, using refined Ogura, Can(napus) and Tour sterile cytoplasms was studied to understand the morphometric and molecular changes. The established Ogura-based CMS line of ‘84 series’ (in the nuclear background of snowball cauliflower) was used as an immediate source for conversion of fertile early- and mid-maturity groups lines. Observations on 18 agro-morphological traits revealed significant differences among the CMS lines, however they could not distinguish different CMS systems. The CMS system significantly reduced the floral traits (19) in almost all the transformed lines irrespective of the maturity group, however, the reduction was maximum in CMS line Tour(CMS 394–41–5) having Tour sterile cytoplasm. This CMS line had a peculiar feature of ‘gripping phenomena’ for the style which drastically reduced flower size and eventually seed yield. These CMS lines showed variation with 11 mtDNA markers and 36 genomic simple sequence repeats (gSSR), and of these, BnTR1 perfectly-identified Can(napus) and Tour CMS systems. This is the first detailed report on the extent of diversity observed in 24 new CMS lines developed using three diverse sterile cytoplasms in the nuclear background of different maturity groups of Indian cauliflower. The information will be of immense significance for immediate use in hybrid breeding and/or use as an immediate source of CMS trait for tropical cauliflowers in other regions.

Cytoplasmic effects of CMS-D2 in upland cotton

<p indent="0mm">Two sets of isonuclear alloplasmic near-isogenic restorer lines of <italic>Gossypium harknessii</italic> were planted in cotton production areas of the Yellow River (Anyang, Henan) and Yangtze River (Jiujiang, Jiangxi) basins, and comparative analyses of seedling growth, leaf photosynthetic parameters, anther development, cotton yield as well as its constituent factors, and fiber quality traits were performed to clarify the cytoplasmic effects of CMS-D2 in upland cotton. The results are as follows: (i) sterile cytoplasm did not affect cotton seedling growth and most fiber quality traits. (ii) The net photosynthetic rate of the sterile cytoplasmic restorer lines was significantly higher at the seedling stage but lower after the blooming stage when compared to the fertile cytoplasmic restorer lines. (iii) The anther fertility of the sterile cytoplasmic restorer lines under high-temperature stress was significantly lower than that of the fertile cytoplasmic restorer lines, which was mainly manifested in a significant reduction in pollen amount, pollen activity and vitality. (iv) The sterile cytoplasm negatively affected yield traits, which was mainly manifested in a significant reduction in boll number per plant and lint percentage. (v) In general, the negative effects of sterile cytoplasm on net photosynthetic rate, anther development and cotton yield were more obvious in cotton production area of the Yangtze River basin where summer temperatures are higher. This paper systematically explored the effects of CMS-D2 cytoplasm on the main traits of upland cotton, and the results of this study are of great significance for the breeding and improvement of cytoplasmic sterile “three-line” hybrid cotton.