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The interaction of plant mitochondrial and nuclear genetic systems is exemplified by mitochondria-encoded cytoplasmic male sterility (CMS) under the control of nuclear restorer-of-fertility genes. The S type of CMS in maize is characterized by a pollen collapse phenotype and a unique paradigm for fertility restoration in which numerous nuclear restorer-of-fertility lethal mutations rescue pollen function but condition homozygous-lethal seed phenotypes. Two nonallelic restorer mutations recovered from Mutator transposon-active lines were investigated to determine the mechanisms of pollen fertility restoration and seed lethality. Mu Illumina sequencing of transposon-flanking regions identified insertion alleles of nuclear genes encoding mitochondrial ribosomal proteins RPL6 and RPL14 as candidate restorer-of-fertility lethal mutations. Both candidates were associated with lowered abundance of mitochondria-encoded proteins in developing maize pollen, and the rpl14 mutant candidate was confirmed by independent insertion alleles. While the restored pollen functioned despite reduced accumulation of mitochondrial respiratory proteins, normal-cytoplasm plants heterozygous for the mutant alleles showed a significant pollen transmission bias in favor of the nonmutant Rpl6 and Rpl14 alleles. CMS-S fertility restoration affords a unique forward genetic approach to investigate the mitochondrial requirements for, and contributions to, pollen and seed development. [ABSTRACT FROM AUTHOR]

Cytoplasmic male sterility (CMS) is a very important factor to produce hybrid seeds, and the restoration of fertility involves the expression of many fertility‐related genes. Our previous study showed that the expression of CaPIPLC5 was significantly up‐regulated in pepper restorer accessions and minimally expressed in sterile accessions, speculating that CaPIPLC5 is related to the restoration of fertility. In this study, we further validated the function of CaPIPLC5 in the restoration of fertility. The results showed that CaPIPLC5 was specifically expressed in the anthers of the restorer accessions with the subcellular localization in the cytoplasm. Furthermore, the expression of CaPIPLC5 was significantly higher in restorer lines and restorer combinations than that in CMS lines and their maintainer lines. Silencing CaPIPLC5 led to the number of pollen decreased, pollen grains wrinkled, and the ratio of pollen germination reduced. In addition, the joint analysis of Yeast One‐Hybrid (Y1H) and Dual‐Luciferase (dual‐LUC) assays suggested that transcription factors such as CaARF5, CabZIP24 and CaMYB‐like1, interacted with the promoter regions of CaPIPLC5, which regulated the expression of CaPIPLC5. The present results provide new insights into the study of CaPIPLC5 involved in the restoration of fertility in pepper. [ABSTRACT FROM AUTHOR]

The cytoplasm of Aegilops kotschyi is known for the induction of male sterility and haploidy in wheat. Both systems originally appeared rather simple, but manipulation of the standard chromosome constitution of the nuclear genome revealed additional interactions. This study shows that while there is little or no allelic variation at the main fertility restorer locus Rfmulti on chromosome arm 1BS, additional genes may also be involved in the nuclear–mitochondrial genome interactions, affecting not only male fertility but also the growth rate, from pollen competition for fertilization and early endosperm divisions all the way to seed size and plant maturity. Some of these effects appear to be of a sporophytic nature; others are gametophytic. Induction of parthenogenesis by a rye inducer in conjunction with the Ae. kotschyi cytoplasm is well known. However, here we show that the cytoplasmic-nuclear interactions affect all aspects of double fertilization: producing maternal haploids from unfertilized eggs, diploids from fertilized eggs or synergids, embryo-less kernels, and fertilized eggs without fertilization of the double nucleus in the embryo sack. It is unclear how frequent the inducers of parthenogenesis are, as variation, if any, is obscured by suppressors present in the wheat genome. Genetic dissection of a single wheat accession revealed five distinct loci affecting the rate of maternal haploid production: four acting as suppressors and one as an enhancer. Only when the suppressing haplotypes are confirmed may it be possible to the identify genetic variation of haploidy inducers, map their position(s), and determine their nature and the mode of action. [ABSTRACT FROM AUTHOR]

Background: As a vital type of noncoding RNAs, circular RNAs (circRNAs) play important roles in plant growth and development and stress response. However, little is known about the biological roles of circRNAs in regulating the stability of male fertility restoration for cytoplasmic male sterility (CMS) conditioned by Gossypium harknessii cytoplasm (CMS-D2) cotton under high-temperature (HT) stress. Results: In this study, RNA-sequencing and bioinformatics analysis were performed on pollen grains of isonuclear alloplasmic near-isogenic restorer lines NH [N(Rf1rf1)] and SH [S(Rf1rf1)] with obvious differences in fertility stability under HT stress at two environments. A total of 967 circRNAs were identified, with 250 differentially expressed under HT stress. We confirmed the back-splicing sites of eight selected circRNAs using divergent primers and Sanger sequencing. Tissue-specific expression patterns of five differentially expressed circRNAs (DECs) were also verified by RT-PCR and qRT-PCR. Functional enrichment and metabolic pathway analysis revealed that the parental genes of DECs were significantly enriched in fertility-related biological processes such as pollen tube guidance and cell wall organization, as well as the Pentose and glucuronate interconversions, Steroid biosynthesis, and N-Glycan biosynthesis pathways. Moreover, we also constructed a putative circRNA-mediated competing endogenous RNA (ceRNA) network consisting of 21 DECs, eight predicted circRNA-binding miRNAs, and their corresponding 22 mRNA targets, especially the two ceRNA modules circRNA346-miR159a-MYB33 and circRNA484-miR319e-MYB33, which might play important biological roles in regulating pollen fertility stability of cotton CMS-D2 restorer line under HT stress. Conclusions: Through systematic analysis of the abundance, characteristics and expression patterns of circRNAs, as well as the potential functions of their parent genes, our findings suggested that circRNAs and their mediated ceRNA networks acted vital biological roles in cotton pollen development, and might be also essential regulators for fertility stability of CMS-D2 restorer line under heat stress. This study will open a new door for further unlocking complex regulatory mechanisms underpinning the fertility restoration stability for CMS-D2 in cotton. [ABSTRACT FROM AUTHOR]

The improvement of grain yield, quality, and resistance can be achieved through the utilization of heterosis. The combination of cytoplasmic male sterility (CMS) and fertility restoration (Rf) gene(s) greatly facilitates the commercial development of three-line hybrid rice based on heterosis. The basis for investigating the relationship between CMS and Rf genes lies in the rapid localization of wild rice fertility restoration genes. A set of the BC4F5 population derived from interspecific crosses between Xieqingzao B (XB) and the BC1F9 XB//Dongxiang wild rice (DWR)/XB line L5339 was used to detect quantitative trait loci (QTL) for fertility restoration. The population was then crossed with two male sterile lines, Zhong9A (Z9A) and DongB11A (DB11A), in order to generate a testcrossing population for investigating spikelet fertility. Based on the linkage mapping, seven QTLs were detected on chromosomes 1, 3, 5, 6, 8, and 10, explaining 2.76 to 12.46% of the phenotypic variation. Of them, two novel fertility restoration QTLs, qRf3 and qRf6, can restore fertility of the CMS-DWR line DB11A by 16.56% and 15.12%, respectively. By employing joint QTL-seq and GradedPool-Seq methods, two novel Rf QTLs for DB11A, qRf3 and qRf6, were identified at the physical locations of 10,900,001–11,700,000 bp and 28,016,785–31,247,556 bp, respectively. These findings are useful for exploring the natural variations of Rf genes in rice. Therefore, rice's new genetic resources for the selection and breeding of rice restorer lines provide promising candidates for QTL fine localization and clarification. [ABSTRACT FROM AUTHOR]

The cellulose synthase gene superfamily contains cellulose synthase (CesA) and cellulose synthase-like (Csl) gene families, which synthesize cellulose and hemicellulose in plant cell walls and play a crucial role in plant growth and development. However, the CesA/Csl gene family has not been reported in pepper. Therefore, the genome-wide research of the CaCesA/CaCsl gene family was conducted in pepper. In this study, a total of 39 CaCesA/CaCsls genes (10 CesAs genes and 29 Csls genes) were identified in pepper and unevenly distributed on 11 chromosomes. These CaCesA/Csls were divided into seven subfamilies (CesAs, CslAs, CslBs, CslCs, CslDs, CslEs, CslGs), and most of CaCesA/Csls genes are closely related to AtCesA/Csls genes. The cis-acting elements in the promoters of CaCesA/Csls genes are mainly related to hormone response and stress response. There are ten collinear gene pairs between the CesA/Csls gene family of pepper and Arabidopsis, and four fragment duplication gene pairs of the CaCesA/Csls genes were discovered. RNA-seq analysis shows that the majority of CaCesA/Csls are expressed in a variety of plant tissues, indicating that most CaCesA/Csls gene expression patterns are not organ-specific, and CaCslD1/D4 have the highest expression in anthers, followed by petal, ovary, and F9. RNA-seq analysis shows that most CaCesA/Csls are responsive to five hormones (IAA, GA3, ABA, SA, and MeJA). The tissue-specific expression analysis of the CaCslD1 gene shows that the CaCslD1 gene is expressed specifically in flowers. In the flower buds IV of cytoplasmic male sterility (CMS) and its restoration of fertility (Rf) system, CaCslD1 reach the highest expression respectively. However, the relative expression level of CaCslD1 in the fertile accessions is extremely significantly higher than in the sterile accessions. This study shows an overall understanding of the CaCesA/Csls gene family and provides a new insight for understanding the function of CaCslD1 in pollen development and exploring the fertility restoration of CMS in pepper. [ABSTRACT FROM AUTHOR]

Cytoplasmic male sterility (CMS) in potato is a common reproductive issue in late blight breeding programs since resistant sources usually have a wild cytoplasmic background (W or D). Nevertheless, in each breeding cycle male fertile lines have been observed within D- and T-type cytoplasms, indicating the presence of a fertility restorer (Rf) mechanism. Identifying sources of Rf and complete male sterility to implement a CMS–Rf system in potato is important since hybrid breeding is a feasible breeding strategy for potato. The objective of this study was to identify male fertile breeding lines and potential Rf candidate lines in the CIP late blight breeding pipeline. We characterized male fertility/sterility-related traits on 142 breeding lines of known cytoplasmic type. We found that pollen viability is not a reliable estimate of male sterility in diverse backgrounds. Breeding lines of the T-type cytoplasmic group had higher levels of male fertility than breeding lines of the D-type cytoplasmic group. With the help of pedigree records, reproductive traits evaluations and test crosses with female clones of diverse background, we identified four male parental lines segregating for Rf and three female parental lines that generated 100% male sterile progeny. These identified lines and generated test cross progenies will be valuable to develop validation populations for mitochondrial or nuclear markers for the CMS trait and for dihaploid generation of Rf+ lines that can be later employed in diploid hybrid breeding. [ABSTRACT FROM AUTHOR]

Rf4 is one of the dominant restorer genes for maize C-type cytoplasmic male sterility (CMS-C), which has significant value in hybrid maize seed production. However, the highly complex fertility restoration mechanism of CMS-C makes it difficult to screen Rf4-restorer lines, and insufficient Rf4-restorer lines limit its use in current agricultural production. To search for Rf4-restorer lines, in this study, the genotypes of eighteen inbred maize lines at the Rf4 locus were analyzed based on the male fertility investigation of hybrid F1, the genetic analysis of F2 populations, molecular marker mapping, allelic tests, and Rf4 genomic sequence analysis. Our results indicated that of the eighteen maize inbred lines, ten were able to completely rescue CMS-C line CHuangzaosi (CHZS) male sterility. A genetic analysis showed that DAN598, PHT77, 78551S, and LH212Ht only contained one dominant restorer gene each, and the molecular-marker mapping indicated that their restorer genes were located at the short arm of chromosome 8. The allelic testing of the fertility of the restorer (Rf) demonstrated that the restorer gene of twelve inbred lines, including DAN598, PHT77, 78551S, and LH212Ht, was allelic to one restorer gene of A619. Furthermore, the genomic sequence alignment of Rf4 revealed that there were two different amino acids in the coding sequence between the A619 (Rf4Rf4) restorer lines and four CMS-C lines (rf4rf4). For the crucial S1596 site variation (TTT/TAC), DAN598, PHT77, 78551S, and LH212Ht shared the same bases (TTT) with A619 and encoded phenylalanine, while the four CMS-C sterile lines had the TAC and encoded tyrosine. Our results revealed that these tester lines, DAN598, PHT77, 78551S, and LH212Ht, were the Rf4-restorer lines. Additionally, derived from the sequence variants of Rf4, 39 possible Rf4-restorer lines from 129 inbred maize lines were detected. Furthermore, we developed a Cleaved Amplified Polymorphism Sequences (CAPS) marker based on the S1596 variations. The PCR amplification product of S1596 (TAC) was digested by the TatI endonuclease into two bands with sizes of ~260 bp and ~100 bp. In comparison, when S1596 was TTT, the PCR product could not be digested. In conclusion, in this study, we identified various Rf4-restorer lines for maize CMS-C and developed a molecular marker for Rf4. The reported results will contribute to the popularization and application of Rf4 in hybrid maize-seed production. [ABSTRACT FROM AUTHOR]

Male‐sterile plants are used in hybrid breeding to improve yield in soybean (Glycine max (L.) Merr.). Developing the capability to alter fertility under different environmental conditions could broaden germplasm resources and simplify hybrid production. However, molecular mechanisms potentially underlying such a system in soybean were unclear. Here, using positional cloning, we identified a gene, MALE STERILITY 3 (MS3), which encodes a nuclear‐localized protein containing a plant homeodomain (PHD)‐finger domain. A spontaneous mutation in ms3 causing premature termination of MS3 translation and partial loss of the PHD‐finger. Transgenetic analysis indicated that MS3 knockout resulted in nonfunctional pollen and no self‐pollinated pods, and RNA‐seq analysis revealed that MS3 affects the expression of genes associated with carbohydrate metabolism. Strikingly, the fertility of mutant ms3 can restore under long‐d conditions. The mutant could thus be used to create a new, more stable photoperiod‐sensitive genic male sterility line for two‐line hybrid seed production, with significant impact on hybrid breeding and production. [ABSTRACT FROM AUTHOR]

Photoperiod/temperature‐sensitive genic male sterility (P/TGMS) is widely applied for improving crop production. Previous investigations using the reversible male sterile (rvms) mutant showed that slow development is a general mechanism for restoring fertility to P/TGMS lines in Arabidopsis. In this work, we isolated a restorer of rvms–2 (res3), as the male sterility of rvms–2 was rescued by res3. Phenotype analysis and molecular cloning show that a point mutation in UPEX1 l in res3 leads to delayed secretion of callase A6 from the tapetum to the locule and tetrad callose wall degradation. Electrophoretic mobility shift assay and chromatin immunoprecipitation analysis demonstrated that the tapetal transcription factor ABORTED MICROSPORES directly regulates UPEX1 expression, revealing a pathway for tapetum secretory function. Early degradation of the callose wall in the transgenic line eliminated the fertility restoration effect of res3. The fertility of multiple known P/TGMS lines with pollen wall defects was also restored by res3. We propose that the remnant callose wall may broadly compensate for the pollen wall defects of P/TGMS lines by providing protection for pollen formation. A cellular mechanism is proposed to explain how slow development restores the fertility of P/TGMS lines in Arabidopsis. [ABSTRACT FROM AUTHOR]

Key message: A candidate gene for male fertility restoration in Brassica juncea, BjRf, was isolated from a 23-kb interval on chromosome A05 using map-based cloning and BSA methods. The cytoplasmic male sterility/fertility restoration (CMS/Rf) system has been extensively used for heterosis in plants. It also provides valuable resources for studying mitochondrial–nuclear coevolution and interaction. The oxa CMS, which is a new CMS type reported in Brassica juncea (B. juncea), has been broadly used in the exploitation and application of heterosis in this species. However, the oxa CMS fertility restorer gene BjRf has not been reported. In this study, a stable restorer line was successfully constructed via continuous testcross and artificial selection. Besides, a new Rf gene was mapped in a 23-kb region on chromosome A05 in B. juncea with a genetic distance of 0.5 cM by the method incorporating bulk segregant analysis (BSA) and conventional map-based cloning. Finally, BjuA017917, a non-PPR Rf gene encoding a guanosine nucleotide diphosphate dissociation inhibitor (GDI), is proposed to be the candidate gene for fertility restoration of the oxa CMS line in B. juncea. Moreover, a functional marker, CRY3, was developed for marker-assisted selection for Brassica juncea breeding. [ABSTRACT FROM AUTHOR]

Number of spikelets per panicle is one of the important characters for rice breeding programs to increase yield potential. The generation mean analysis based on six populations: viz., P1, P2, F1, F2, BC1, and BC2 of IR80151A/CH1 with utilizing cytoplasmic-genetic male, was performed. The results showed that dominant gene action was more important than additive type implying that selection in early segregating generations is ineffective and the character could be better exploited through heterosis breeding. Since hybrid rice was used the advantage of the dominant effect through heterosis, due to the role of more favorable dominant genes in F1 hybrid than in both parents. In addition, utilizing cytoplasmic-genetic male sterility in the populations could specify gene control of fertility restoration in hybrid rice. The results revealed that two dominant complementary genes with epistasis controlled the fertility-restoration of wild-abortive cytoplasmic male sterility system (WA-CMS). This information will be useful for future rice breeding in three-line hybrid system. [ABSTRACT FROM AUTHOR]

Cytoplasmic male sterility (CMS) is extensively used in maize hybrid production, and identification of genes related to fertility restoration for CMS is important for hybrid breeding. The fertility restoration of S type CMS is governed by several loci with major and minor effects, while the mechanism of fertility restoration for CMS-S is still unknown. In this study, BSR-Seq was conducted with two backcrossing populations with the fertility restoration genes, Rf3 and Rf10, respectively. Genetic mapping via BSR-Seq verified the positions of the two loci. A total of 353 and 176 differentially expressed genes (DEGs) between the male fertility and male sterile pools were identified in the populations with Rf3 and Rf10, respectively. In total, 265 DEGs were co-expressed in the two populations, which were up-regulated in the fertile plants, and they might be related to male fertility involving in anther or pollen development. Moreover, 35 and seven DEGs were specifically up-regulated in the fertile plants of the population with Rf3 and Rf10, respectively. Function analysis of these DEGs revealed that jasmonic acid (JA) signal pathway might be involved in the Rf3 mediated fertility restoration for CMS-S, while the small ubiquitin-related modifier system could play a role in the fertility restoration of Rf10. [ABSTRACT FROM AUTHOR]

Thirty‐one tropical japonica derived Rf gene carrying rice hybrids were classified into three classes as Rf3, Rf4 and Rf3 + Rf4 hybrids. These hybrids were tested under three different mega‐environments. Between Rf3 class and both the classes of hybrids possessing Rf4 genes, significant variation for spikelet and pollen fertility and grain yield was found. The pollen fertility was five times higher among Rf4 hybrids than that of hybrids carrying Rf3 alone. Likewise, spikelet fertility among Rf4 hybrids was two times higher than that of Rf3 hybrids. Parallel grain yield increase in Rf4 hybrids was 2.4 times than in Rf3 hybrids. However, Rf3 gene was found complementing Rf4 to truncate the range of pollen and spikelet fertility. Hybrids possessing Rf3 alone exhibited partial pollen and spikelet sterility, and significant negative standard heterosis for grain yield. The high yielding hybrid, "HYB36" carrying both the Rf genes was found to be widely adapted. The present study established that Rf4 gene is essential either alone or in combination with Rf3 for fertility restoration to achieve enhanced grain yield in WA‐CMS based hybrids. [ABSTRACT FROM AUTHOR]

Inheritance of fertility restorer gene in pigeonpea was studied using F2 and BC1F1 populations derived from cross AL103A × IC245273. It was found to be controlled by single dominant gene. Out of 228 SSR primer pairs, 33 primer pairs showed parental polymorphism, while four primers were found polymorphic in bulk segregant analysis (BSA). These four primers viz., CcM 1615, CcM 0710, CcM 0765 and CcM 1522 were used for genotyping of F2 population and were found to be placed at 3.1, 5.1, 28.1 and 45.8 cM, respectively. Two of them, CcM 1615 and CcM 0710, evinced clear and unambiguous bands for fertility restoration in F2 population. The Rf gene was mapped on linkage group 6 between the SSR markers CcM 1615 and CcM 0710 with the distances of 3.1 and 5.1 cM, respectively. The accuracy of the CcM 1615 was validated in 18 restorers and six maintainer lines. The marker CcM 1615 amplified in majority of male restorer lines with a selection accuracy of 91.66%. [ABSTRACT FROM AUTHOR]

The progenies from crosses of a sterile VIR116A line with PET1 CMS with pollen fertility restorer RIL130, VIR210, and VIR740 lines, differing in the type of cytoplasm and the presence of diagnostic molecular markers of the Rf1 gene, were studied. The pollen of F1 plants of all three cross combinations was highly fertile (89.9–98.3% of stained pollen grains, PG). Each of the F2 populations (VIR116A × RIL130, VIR116A × VIR210, and VIR116A × VIR740) segregated for the pollen fertility trait in a ratio 3 (fertile) : 1 (sterile). Among fertile F2 genotypes, "low pollen" ones were identified, in which there was less pollen than normal during a visual estimation. Their frequency was approximately 9% of the total number of plants in each of the populations, which is consistent with the results obtained previously during the analysis of other hybrid combinations. In cytological analysis, the pollen of "low pollen" plants was characterized by a decreased (as compared with highly fertile plants) frequency of fertile PG (80–100 and 11–78%, respectively) and higher content of anomalous PG (in "low pollen" plants up to 88.5%, in plants with normal level of fertility up to 18%). Using HRG01 and HRG02 molecular markers, the presence of the Rf1 dominant allele in the genotypes of fertile F2 plants (including "low pollen" ones) was confirmed, indicating their high diagnostic value. Possible genetic mechanisms of the occurrence of "low pollen" F2 segregants are discussed. [ABSTRACT FROM AUTHOR]

Hybrid rice based on wild‐abortive cytoplasmic male sterility (WA‐CMS) is important in boosting rice production, which requires diverse parents to harness heterosis. For this, exploiting the diversity of japonica through tropical japonica (TRJ) lines is an excellent route. In this study, 310 TRJ‐based new plant type (NPT) lines were developed and evaluated for Rf3 and Rf4 genes. Gene‐based (DRRM‐Rf3‐5 and DRRM‐Rf3‐10) and functional marker (RMS‐SF21‐5) targeted Rf3 locus, while gene‐linked (RM6100) and functional marker (RMS‐PPR9‐1) targeted the Rf4 locus. The frequency of the restorer allele of Rf3 gene was lower when compared to that of Rf4. Combined phenotypic and molecular screening using gene‐based and functional markers identified 42 lines that carried Rf3 and/or Rf4 genes. All the selected lines produced fertile F1s when crossed to a WA‐CMS line, "Pusa 6A", but with varying levels of spikelet fertility. This is the first report of a marker‐cum‐phenotype‐based restorer selection using TRJ‐derived lines. Multilocation evaluation of these lines at three locations indicated better adaptation for grain yield in some of the lines. [ABSTRACT FROM AUTHOR]

In order to introduce the Tomato Spotted Wilt Virus ( TSWV) resistance from Nicotiana alata into Nicotiana tabacum, a cytoplasmic male sterility ( CMS) line of N. tabacum ( N. tabacum L. cv. ( gla.) S 'K326'), was successfully crossed with N. alata. Despite a high DNA content variability, F1 hybrids could be classified in two subgroups, a major one encompassing fertile hybrids morphologically similar to their tobacco maternal parent but TSWV sensitive, and a minor one displaying sterile hybrids showing an intermediate phenotype and TSWV resistant. In order to elucidate the unexpected fertility recovery of the fertile F1 plants, some N. alata fertility restoration ppr genes were cloned and were shown to be differentially expressed between parental lineages as well as between both F1 subgroups, suggesting that N. alata contains fertility restoring allele able to overcome the CMS of N. tabacum. [ABSTRACT FROM AUTHOR]

RPHR-1005, the stable restorer line of the popular medium slender (MS) grain type rice hybrid, DRRH-3 was improved in this study for resistance against bacterial blight (BB) and blast diseases through marker-assisted backcross breeding (MABB). In this study, four major resistance genes (i.e., Xa21 and Xa33 for BB resistance and Pi2 and Pi54 for blast resistance) have been transferred to RPHR-1005 using RPBio Patho-1 (possessing Xa21 + Pi2), RPBio Patho-2 (possessing Xa21 + Pi54) and FBR1-15EM (possessing Xa33) as the donors. Foreground selection was carried out using PCR-based molecular markers specific for the target resistance genes and the major fertility restorer genes, Rf3 and Rf4, while background selection was carried out using a set of parental polymorphic rice SSR markers and backcrossing was continued uptoBC2 generation. At BC2F2, plants possessing the gene combination- Xa21 + Pi2, Xa21 + Pi54 and Xa33 in homozygous condition and with >92% recovery of the recurrent parent genome (RPG) were identified and intercrossed to combine all the four resistance genes. Twenty-two homozygous, pyramid lines of RPHR-1005 comprising of three single-gene containing lines, six 2-gene containing lines, eight 3-gene containing lines, and five 4-gene containing lines were identified among the double intercross lines at F3 generation (DICF3). They were then evaluated for their resistance against BB and blast, fertility restoration ability and for key agro-morphological traits. While single gene containing lines were resistant to either BB or blast, the 2-gene, 3-gene, and 4-gene pyramid lines showed good level of resistance against both and/or either of the two diseases. Most of the 2-gene, 3-gene, and 4-gene containing pyramid lines showed yield levels and other key agro-morphological and grain quality traits comparable to the original recurrent parent and showed complete fertility restoration ability, with a few showing higher yield as compared to RPHR-1005. Further, the experimental hybrids derived by crossing the gene-pyramid lines of RPHR-1005 with APMS6A (the female parent of DRRH-3), showed heterosis levels equivalent to or higher than DRRH-3. The results of present study exemplify the utility of MABB for targeted improvement of multiple traits in hybrid rice. [ABSTRACT FROM AUTHOR]

A total of sixty-six germplasm lines were crossed with five CMS lines, where two belong to A4 cytoplasm, while other three belong to A2 cytoplasm. On the basis of pollen fertility test as well as good pod setting, of 330 hybrids, 34 restorer lines were observed in ICPA 2043 and 19 in ICPA 2092. Thirteen germplasm lines restored fertility in both the A4 CMS lines, viz. ICPA 2043 and ICPA 2092; however, none of the lines restored fertility in A2 CMS lines. For confirmation of result, restoration competence of identified lines tested subsequently 2 years at two different temperatures. The segregation patterns for fertility restoration studied in F2 and BC1F1 generations of selected ten crosses. Six crosses indicated the involvement of two major genes with recessive epistasis, three crosses confirmed dominant epistasis, and one cross indicated the involvement of duplicate recessive epistasis. The obtained results from this study will hasten the future three-line breeding programme and lead the hybrid technology to the farmers' field with the better exploitation of CMS lines. [ABSTRACT FROM AUTHOR]

The PPR-B gene is responsible for male-fertility restoration of the Ogura-type male-sterile radish plants, and it is located in the complex Rfo locus in the vicinity of similar PPR-A gene and PPR-C pseudogene. The aim of this study was to identify PPR-B alleles and understand the structure of the Rfo locus in radish breeding lines. Five lines of radish with normal male-fertile cytoplasm were tested. The entire PPR-B gene was amplified, sequenced and allelic PPR-B sequences were identified. The results indicated that the maintainer lines 7, 15 and 21 contained a non-restoring form of PPR-B protein. A unique PPR-B was found in lines 24/15 and 31 that are restorer and maintainer lines, respectively. The substitutions might be responsible for the loss of a restoring function of the PPR-B-31 allele. Amplification of the PPR-A/ PPR-B and PPR-B/ PPR-C intergenic regions allowed to identify rearrangements within Rfo locus. Obtained results confirm the wide allelic variation within the Rfo locus, as well as high genetic complexity of the fertility restoration mechanism in radish. [ABSTRACT FROM AUTHOR]

Investigation was carried out to identify restorers/maintainers with WA based CMS lines. Various inbred lines behaved differently in four CMS back grounds in respect of their maintainer/restorer reaction. Lines R-42, R-43, R-44, R-46, R-47, R-56, R-51, R-52, R-54, IR-64, IR-66, DRR-714, IR10198, IR 13419 and JGL 11118 identified as potential restorers at over the locations and these lines can be utilized for developing new potential hybrids. Fertility restoration studies indicated that restorer lines (R-43, R-44, R-52 and R-54) were governed by two independent dominant genes. A restorer with more than one independent gene is likely to produce the hybrids with higher fertility restoration and consequently increased yield. [ABSTRACT FROM AUTHOR]

Segregating patterns for pollen fertility of five crosses were studied involving two CGMS lines and four restorers viz., ICPA 2043/ICP 6399, ICPA 2043/ICP 9149, ICPA 2043/ICPR 4105, ICPA 2092/ICPR 4105 and ICPA 2092/KA 91-25 in F2 and BC1F1 generations. The crosses exhibited 13: 3 and 3:1 ratios, respectively in F2 and BC1F1 indicating the involvement of two dominant genes with one basic and one inhibitory gene action for fertility restoration. However, three crosses, ICPA 2092/ICP 6399, ICPA 2092/ICP 9149 and ICPA 2043/KA 91-25 recorded the ratio of 9 : 3 : 4, 12 : 3 : 1 and 9 : 7 and 1 : 1 : 2, 2 : 1 : 1 and1 : 3 in F2 and BC1F1 populations, respectively and these are confirming that supplementary, masking and complimentary gene actions. [ABSTRACT FROM AUTHOR]

Cytoplasmic male sterility ( CMS) hybrid rice has made a great contribution to the increase of rice yield globally. To facilitate the development of high-quality pairs of the wild abortive ( WA) male sterile and maintainer lines, the genetic basis of fertility restoration of WA- CMS and stigma exsertion was investigated in this study using a testcross population with the WA- CMS background. Seed-setting rate and stigma exsertion rate were used as the indicators of the two traits, respectively. Results showed that four minor QTL regions from 9311 were responsible for the variation of seed-setting rate, while a few minor QTLs and epistatic QTL pairs influenced stigma exsertion rate. These results would be of great use in the development of high-quality pairs of WA male sterile and maintainer lines in rice. [ABSTRACT FROM AUTHOR]

The success of hybrid rice technology depends on the extent of heterosis realized, pest resistance and the grain quality of rice. To identify gall midge resistant, non aromatic maintainers and restorers, 114 germplasm lines were crossed with six CMS lines (2 indigenous and four exotic) to get one hundred fifty five hybrids (Rabi, 2012-13) and evaluated with 10 checks (Kharif, 2013) at Rice Research Station, Regional Agricultural Research Station, Jagtial, Andhra Pradesh. Based on pollen sterility and spikelet fertility studies, 49 maintainers and 31 restorers were identified. Screening hybrids and their parental lines for gall midge incidence indicated the involvement of dominant genes for gall midge resistance. Forty three hybrids, eleven maintainers and eighteen restorers were resistant to gall midge biotype 3. Gall midge resistant maintainers and restorers will be used for new CMS line and hybrid development respectively. The conventional method of restore line identification among rice germplasm pool is time consuming and labor intensive. Molecular mapping of fertility restorer genes in rice have yielded several closely linked DNA markers that can be used in identifying restorer lines. In order to utilize this available information effectively in marker assisted restorer line identification, validation of reported Rf3 and Rf4 gene linked DNA markers was carried out in this study. A total of seven DNA markers reported to be closely linked with two Rf genes of wild abortive CMS (WA-cms) were chosen. These markers were screened among twenty identified restorers and five maintainer lines. The genotypic data set was generated based on the specific PCR product size. Two DNA markers in combination (RM10313 and RM6100) showed 100% selection efficiency in identifying restorers in the germplasm and 90% selection efficiency in differentiating maintainers from restorers. These validated molecular markers linked to Rf genes would save time and money besides adding accuracy in identification of restorers. [ABSTRACT FROM AUTHOR]

Highlights: [•] Fertility restoration by the Rf genes in three CMS systems of eggplant was studied. [•] Each Rf gene restores fertility of other CMS systems as well as its own CMS system. [•] RAPD markers linked to the Rf genes were screened by bulked segregant analysis. [•] One Operon primer OPAB101900 was identified as linked to the Rf genes. [•] The RAPD marker was converted to SCAR marker designated as SCAB101900. [Copyright &y& Elsevier]

The molecular and biochemical bases of fertility restoration were explored using cytoplasmic male sterile (CMS) Ji A, maintainer Ji B, and two cotton hybrids RF and QF, developed by crossing CMS with DES-HAF277 (normal restorer) and Zheda strong restorer (transgenic restorer with GST gene), respectively. Transcript levels of both exogenous and endogenous GST genes were high in anther as compared to other plant tissues of the QF hybrid. Moreover, the expression of the GST gene during meiosis (stage 2) and microspore development (stage 3) was highest in the QF hybrid. The ratio of cyanide-resistant respiration to total respiration was also high in the QF hybrid during stage 2 and stage 3 as compared to the RF hybrid. O and HO contents increased more during stage 2 in the CMS line and stage 3 in the RF hybrid compared to the maintainer and QF hybrid. Similarly, MDA contents were at a maximum in CMS followed by the RF, QF, and the maintainer line during the whole course of anther development. In addition, the activities of antioxidant enzymes (SOD, POD, CAT, APX, GST, GR, and DHAR) and contents of non-enzymatic antioxidants (GSH and ASA) were elevated in the QF hybrid as compared to the RF hybrid, during the whole course of anther development. The present study suggests that the introgression of the GST gene into restorer lines could be a potential way to enhance restoration capability by maintaining the equilibrium between oxidative stress and scavenging enzymes, and might favor healthier development of microspores. [ABSTRACT FROM AUTHOR]

Abstract: Restorer line breeding is an important approach to enhance the heterosis and improve the yields of japonica hybrid rice. To improve the selection efficiency of restorer lines for BT-type cytoplasmic male sterility (CMS) in japonica rice, a functional marker InDel-Rf-1a based on the difference of nucleotide sequence in Rf-1a locus between BT-type CMS lines and restorer lines was developed to detect the genotypes of different rice materials. Conventional indica rice varieties, restorer and maintainer lines without 574 bp deletion could restore the fertility for BT-type CMS in japonica rice. By contrast, most conventional japonica rice varieties except Aichi 106 and Yijing 12, with genotype of rf-1arf-1a showed the 574 bp deletion maintained sterility for BT-type CMS lines. To further verify the effect of genotyping detection in Rf-1a locus, this marker was also used to amplify the genomic DNA in different japonica rice restorer lines, CMS lines, hybrids and F2 segregation population, and three genotypes in Rf-1a locus could be distinguished distinctly. Therefore, the marker InDel-Rf-1a could be widely used for genetic identification and marker-assisted selection (MAS) in breeding japonica restorer lines. [Copyright &y& Elsevier]

Among the natural mechanisms used for wheat hybrid breeding, the most desirable is the system combining the cytoplasmic male sterility (cms) of the female parent with the fertility-restoring genes (Rf) of the male parent. The objective of this study was to identify Rf candidate genes in the wheat genome on the basis of transcriptome sequencing (RNA-seq) and paralog analysis data. Total RNA was isolated from the anthers of two fertility-restorer (Primépi and Patras) and two non-restorer (Astoria and Grana) varieties at the tetrad and late uninucleate microspore stages. Of 36,912 differentially expressed genes (DEGs), 21 encoding domains in known fertility-restoring proteins were selected. To enrich the pool of Rf candidates, 52 paralogs (PAGs) of the 21 selected DEGs were included in the analyses. The expression profiles of most of the DEGs and PAGs determined bioinformatically were as expected (i.e., they were overexpressed in at least one fertility-restorer variety). However, these results were only partially consistent with the quantitative real-time PCR data. The DEG and PAG promoters included cis-regulatory elements common among PPR-encoding genes. On the basis of the obtained results, we designated seven genes as Rf candidate genes, six of which were identified for the first time in this study. [ABSTRACT FROM AUTHOR]

Infertility is a globally recognized issue caused by different reproductive disorders. To date, various therapeutic approaches to restore fertility have been attempted including etiology-specific medication, hormonal therapies, surgical excisions, and assisted reproductive technologies. Although these approaches produce results, however, fertility restoration is not achieved in all cases. Advances in using stem cell (SC) therapy hold a great promise for treating infertile patients due to their abilities to self-renew, differentiate, and produce different paracrine factors to regenerate the damaged or injured cells and replenish the affected germ cells. Furthermore, SCs secrete extracellular vesicles (EVs) containing biologically active molecules including nucleic acids, lipids, and proteins. EVs are involved in various physiological and pathological processes and show promising non-cellular therapeutic uses to combat infertility. Several studies have indicated that SCs and/or their derived EVs transplantation plays a crucial role in the regeneration of different segments of the reproductive system, oocyte production, and initiation of sperm production. However, available evidence triggers the need to testify the efficacy of SC transplantation or EVs injection in resolving the infertility issues of the human population. In this review, we highlight the recent literature covering the issues of infertility in females and males, with a special focus on the possible treatments by stem cells or their derived EVs. [ABSTRACT FROM AUTHOR]

Two popular stable restorer lines, CB 87 R and CB 174 R, were improved for blast resistance through marker-assisted back-cross breeding (MABB). The hybrid rice development program in South India extensively depends on these two restorer lines. However, these restorer lines are highly susceptible to blast disease. To improve the restorer lines for resistance against blasts, we introgressed the broad-spectrum dominant gene Pi54 into these elite restorer lines through two independent crosses. Foreground selection for Pi54 was done by using gene-specific functional marker, Pi54 MAS, at each back-cross generation. Back-crossing was continued until BC3 and background analysis with seventy polymorphic SSRs covering all the twelve chromosomes to recover the maximum recurrent parent genome was done. At BC3F2, closely linked gene-specific/SSR markers, DRRM-RF3-10, DRCG-RF4-8, and RM 6100, were used for the identification of fertility restoration genes, Rf3 and Rf4, along with target gene (Pi54), respectively, in the segregating population. Subsequently, at BC3F3, plants, homozygous for the Pi54 and fertility restorer genes (Rf3 and Rf4), were evaluated for blast disease resistance under uniform blast nursery (UBN) and pollen fertility status. Stringent phenotypic selection resulted in the identification of nine near-isogenic lines in CB 87 R × B 95 and thirteen in CB 174 R × B 95 as the promising restorer lines possessing blast disease resistance along with restoration ability. The improved lines also showed significant improvement in agronomic traits compared to the recurrent parents. The improved restorer lines developed through the present study are now being utilized in our hybrid development program. [ABSTRACT FROM AUTHOR]

The PET2-cytoplasm represents a well characterized new source of cytoplasmic male sterility (CMS) in sunflower. It is distinct from the PET1-cytoplasm, used worldwide for commercial hybrid breeding, although it was, as PET1, derived from an interspecific cross between Helianthus. petiolaris and H. annuus. Fertility restoration is essential for the use of CMS PET2 in sunflower hybrid breeding. Markers closely linked to the fertility restorer gene are needed to build up a pool of restorer lines. Fertility-restored F1-hybrids RHA 265(PET2) × IH-51 showed pollen viability of 98.2% ± 1.2, indicating a sporophytic mode of fertility restoration. Segregation analyses in the F2-population of the cross RHA 265(PET2) × IH-51 revealed that this cross segregated for one major restorer gene Rf-PET2. Bulked-segregant analyses investigating 256 amplified fragment length polymorphism (AFLP) primer combinations revealed a high degree of polymorphism in this cross. Using a subset of 24 AFLP markers, three sequence-tagged site (STS) markers and three microsatellite markers, Rf-PET2 could be mapped to the distal region of linkage group 13 between ORS1030 and ORS630. Three AFLP markers linked to Rf-PET2 were cloned and sequenced. Homology search against the sunflower genome sequence of HanXRQ v1r1 confirmed the physical location of Rf-PET2 close to the restorer gene Rf1 for CMS PET1. STS markers were mapped that can now be used for marker-assisted selection. [ABSTRACT FROM AUTHOR]

The cytoplasmic male sterility (CMS)/restorer-of-fertility system is an important tool to exploit heterosis during commercially hybrid seed production. The importance of long noncoding RNAs (lncRNAs) in plant development is recognized, but few analyses of lncRNAs during anther development of three-line hybrid cotton (CMS-D2 line A, maintainer line B, restorer-of-fertility line R) have been reported. Here, we performed transcriptome sequencing during anther development in three-line hybrid cotton. A total of 80,695 lncRNAs were identified, in which 43,347 and 44,739 lncRNAs were differentially expressed in A–B and A–R comparisons, respectively. These lncRNAs represent functional candidates involved in CMS and fertility restoration. GO analysis indicated that cellular hormone metabolic processes and oxidation–reduction reaction processes might be involved in CMS, and cellular component morphogenesis and small molecular biosynthetic processes might participate in fertility restoration. Additionally, 63 lncRNAs were identified as putative precursors of 35 miRNAs, and quantitative reverse transcription polymerase chain reaction (qRT-PCR) showed a similar expression pattern to RNA-seq data. Furthermore, construction of lncRNA regulatory networks indicated that several miRNA–lncRNA–mRNA networks might be involved in CMS and fertility restoration. Our findings provide systematic identification of lncRNAs during anther development and lays a solid foundation for the regulatory mechanisms and utilization in hybrid cotton breeding. [ABSTRACT FROM AUTHOR]

The phenomenon of cytoplasmic male sterility (CMS), consisting in the inability to produce functional pollen due to mutations in mitochondrial genome, has been described in more than 150 plant species. With the discovery of nuclear fertility restorer (Rf) genes capable of suppressing the CMS phenotype, it became possible to use the CMS-Rf genetic systems as the basis for practical utilization of heterosis effect in various crops. Seed production of sunflower hybrids all over the world is based on the extensive use of the PET1 CMS combined with the Rf1 gene. At the same time, data on Rf1 localization, sequence, and molecular basis for the CMS PET1 type restoration of fertility remain unknown. Searching for candidate genes of the Rf1 gene has great fundamental and practical value. Therefore, in this study, association mapping of fertility restorer gene for CMS PET1 in sunflower was performed. The genome-wide association study (GWAS) results made it possible to isolate a segment 7.72 Mb in length on chromosome 13, in which 21 candidates for Rf1 fertility restorer gene were identified, including 20 pentatricopeptide repeat (PPR)family genes and one Probable aldehyde dehydrogenase gene. The results will serve as a basis for further study of the genetic nature and molecular mechanisms of pollen fertility restoration in sunflower, as well as for further intensification of sunflower breeding. [ABSTRACT FROM AUTHOR]

Single dominant fertility restoration (Rf) genes for cytoplasmic male-sterile (CMS) sunflower lines CMS ANN2 and CMS ANN3 were previously identified in three and five sources, respectively. However, expressions of the dominant Rf genes were suppressed when crossing restoration lines with three inbred lines. Preliminary results indicated that segregation of fertility in most testcross progeny families from crosses of half-diallel F1's of restoration lines with the two CMS lines, respectively, did not agree with those of the F2 generation of the half-diallel crosses. In this study, using selected CMS ANN2 and CMS ANN3 plants without the silencing suppressor (S) genes, we confirmed the existence of S genes in the nuclear genomes of the CMS lines used for crossing of the two CMS cytoplasms. Inheritance study suggested that Rf gene suppression was controlled by a single dominant gene in specific backgrounds, whereas two to three complementary S genes could exist in certain backgrounds. Using testcross population Rf ANN2-P21/RHA 274//Rf ANN2-P21, a single S gene from RHA 274, designated S1, was mapped to a recombination suppressed region on the sunflower linkage group (LG) 3, co-segregating with 16 SSR or single nucleotide polymorphism (SNP) markers, flanked by NSA_003339 and ORS124 at a distance of 1.4 and 0.7 cM, respectively. These molecular markers will potentially assist in identifying lines without suppressor genes, including CMS lines. Key message: Unique suppression of Rf genes confirmed existence of suppressor (S1) gene in CMS lines that mapped to a recombination suppressed region on linkage group 3, which will enhance understanding of cytoplasm-nuclear interactions. [ABSTRACT FROM AUTHOR]

Background: C-type cytoplasmic male sterility (CMS-C) is one of the three major types of cytoplasmic male sterility (CMS) in maize. Rf4 is a dominant restorer gene for CMS-C and has great value in hybrid maize breeding, but little information concerning its functional mechanism is known. Results: To reveal the functional mechanism of Rf4, we developed a pair of maize near-isogenic lines (NILs) for the Rf4 locus, which included a NIL_rf4 male-sterile line and a NIL_Rf4 male fertility-restored line. Genetic analysis and molecular marker detection indicated that the male fertility of NIL_Rf4 was controlled by Rf4. Whole-genome sequencing demonstrated genomic differences between the two NILs was clustered in the Rf4 mapping region. Unmapped reads of NILs were further assembled to uncover Rf4 candidates. RNA-Seq was then performed for the developing anthers of the NILs to identify critical genes and pathways associated with fertility restoration. A total of 7125 differentially expressed genes (DEGs) were identified. These DEGs were significantly enriched in 242 Gene Ontology (GO) categories, wherein 100 DEGs were involved in pollen tube development, pollen tube growth, pollen development, and gametophyte development. Homology analysis revealed 198 male fertility-related DEGs, and pathway enrichment analysis revealed that 58 DEGs were enriched in cell energy metabolism processes involved in glycolysis, the pentose phosphate pathway, and pyruvate metabolism. By querying the Plant Reactome Pathway database, we found that 14 of the DEGs were involved in the mitochondrial tricarboxylic acid (TCA) cycle and that most of them belonged to the isocitrate dehydrogenase (IDH) and oxoglutarate dehydrogenase (OGDH) enzyme complexes. Transcriptome sequencing and real-time quantitative PCR (qPCR) showed that all the above TCA cycle-related genes were up-regulated in NIL_Rf4. The results of our subsequent enzyme-linked immunosorbent assay (ELISA) experiments pointed out that the contents of both the IDH and OGDH enzymes accumulated more in the spikelets of NIL_Rf4 than in those of NIL_rf4. Conclusion: The present research provides valuable genomic resources for deep insight into the molecular mechanism underlying CMS-C male fertility restoration. Importantly, our results indicated that genes involved in energy metabolism, especially some mitochondrial TCA cycle-related genes, were associated with maize CMS-C male fertility restoration. [ABSTRACT FROM AUTHOR]