Cytoplasmic Sterility Research Articles

Six New Genes Required for Production of T-Toxin, a Polyketide Determinant of High Virulence of Cochliobolus heterostrophus to Maize

Southern Corn Leaf Blight, one of the worst plant disease epidemics in modern history, was caused by Cochliobolus heterostrophus race T, which produces T-toxin, a determinant of high virulence to maize carrying Texas male sterile cytoplasm. The genetics of T-toxin production is complex and the evolutionary origin of associated genes is uncertain. It is known that ability to produce T-toxin requires three genes encoded at two unlinked loci, Tox1A and Tox1B, which map to the breakpoints of a reciprocal translocation. DNA associated with Tox1A and Tox1B sums to about 1.2 Mb of A+T rich, repeated DNA that is not found in less virulent race O or other Cochliobolus species. Here, we describe identification and targeted deletion of six additional genes, three mapping to Tox1A and three to Tox1B. Mutant screens indicate that all six genes are involved in T-toxin production and high virulence to maize. The nine known Tox1 genes encode two polyketide synthases (PKS), one decarboxylase, five dehydrogenases, and one unknown protein. Only two have a similar phylogenetic profile. To trace evolutionary history of one of the core PKS, DNA from more than 100 Dothideomycete species were screened for homologs. An ortholog (60% identity) was confirmed in Didymella zeae-maydis, which produces PM-toxin, a polyketide of similar structure and biological specificity as T-toxin. Only one additional Dothideomycete species, the dung ascomycete Delitschia winteri harbored a paralog. The unresolved evolutionary history and distinctive gene signature of the PKS (fast-evolving, discontinuous taxonomic distribution) leaves open the question of lateral or vertical transmission.

Inheritance of Fertility Restoration for Cytoplasmic Male Sterility in a New Gossypium barbadense Restorer

In order to clarify inheritance mechanism of fertility restoration for cytoplasmic male sterility (CMS) in a new Gossypium barbadense restorer line Hai R which was found in the fertility test crossing of G. hirsutum CMS lines with G. barbadense germplasms. 23 fertility segregation populations of F 2 and backcross were used to analyze the inheritance of fertility restoring gene(s) of Hai R. The result showed that Hai R had one major dominant gene ( Rf B) to control the CMS fertility restoration and this fertility restoration gene functioned at the sporophytic level. The sterile cytoplasm background might not only influence the transmission rate of male gamete but also that of female gamete when the restorer gene was recessive. It could be deduced that this fertility restoration gene might come from G. harknessii cotton, Hai R is of value in the application of cotton interspecific hybrid breeding.

Classification of the male sterile cytoplasms in rice

Classification of the male sterile cytoplasms in rice

Identification of Male Sterile Cytoplasm and Restorer Gene and Allelism of Restorer Gene in Restorer Lines in Rice

Identification of Male Sterile Cytoplasm and Restorer Gene and Allelism of Restorer Gene in Restorer Lines in Rice

Classification of Male Sterile Cytoplasms Derived from Wild Species in Rice

Classification of Male Sterile Cytoplasms Derived from Wild Species in Rice

Classification of male sterile cytoplasms and fertility restoring genes in rice

Classification of male sterile cytoplasms and fertility restoring genes in rice

Factors Causing a Decrease in Spikelet Fertility in the Rice Plants Carrying Male Sterile Cytoplasm with Rfrf Genotype

Factors Causing a Decrease in Spikelet Fertility in the Rice Plants Carrying Male Sterile Cytoplasm with Rfrf Genotype

Comparative Study on Breeding Utilization Characteristics of the Isonuclear Alloplasmic <I>Japonica</I> CMS Lines Liuqianxin A with Four Different Cytoplasm Sources

A set of isonuclear alloplasmic japonica CMS lines Liuqianxin A,constructed with four different cytoplasm sources from Hainan Male Sterile Wild Rice (WA type),Hainan Red Awned Wild Rice (HL type),Hunan Chaling Wild Rice (CL type),and India Chinsurah Boro II (BT type),respectively,together with their maintainer line Liuqianxin B,were applied to study their male sterile stability,flowering habits,outcrossing rate,and restorable ability.The results showed:(1) Most pollen granules of BT,HL and CL Liuqianxin A were mainly stained-abortion,while those of WA Liuqianxin A were mainly typical-abortion.Selfed seed-setting rate of BT Liuqianxin A was 0.60%,which would cause application risk in agricultural production.There were no selfed seeds in HL and CL Liuqianxin A,indicating that sterility of them was better than that of BT type.(2) The length of 1st internode to the top was affected by the degree of abortive pollen,which influenced the plant height and the panicle exsertion (i.e.the length of panicle-neck above the sheath of flag leaf).Namely the more serious abortive degree,the shorter panicle exsertion.Therefore,panicle was enclosed within the sheath of the flag leaf in WA Liuqianxin A,not in BT,HL,and CL Liuqianxin A.(3) The flowering habits of WA Liuqianxin A presented as sporadic flowering,distinctly lagged flowering time and stretched flowering duration,and were the worst in the four type CMS lines,thus its outcrossing rate was lower.Flowering habits and outcrossing rate between HL,CL CMS lines and BT CMS lines were not significantly different.However it was better in HL,CL CMS lines than in WA CMS lines.(4) BT Liuqianxin A showed the best restorable characteristic,HL was a little worse than BT,WA was the worst.And japonica restorers for HL or CL CMS lines could be selected from the japonica restorers for BT CMS lines.HL,CL CMS lines balanced above contradictions well,so an idea of substituting HL,CL CMS lines partly for BT type was put forward,it can be used to solve the problems of single type sterile cytoplasm in current Japonica hybrid rice and fertility not stable in BT CMS lines.

Haplotype analysis of CMS-associated DNA markers in sweet peppers

Cytoplasmic male sterility (CMS)/restorer-of-fertility (Rf) is an economical and efficient system to produce F1 hybrid seeds. Although the CMS/Rf system has been used to produce hybrid seeds of hot peppers, this system has never been used for sweet pepper seed production, presumably due to the inability to select stable restorer lines during the breeding process. To test the feasibility of the CMS/Rf system in sweet pepper breeding, we investigated the distribution of haplotypes of previously developed, CMS-associated markers (orf456, ψ atp6-2, CRF-SCAR, OPP13-CAPS, PR-CAPS, and PR-SNP) in 27 commercial sweet pepper F1 hybrids and 12 breeding lines. When CMS-associated cytoplasmic markers orf456 and ψ atp6-2 were applied, male sterile cytoplasm was not detected in commercial sweet pepper cultivars. When nuclear haplotype markers linked to Rf were applied, all sweet pepper cultivars showed haplotype 3, haplotype 1, and the rf genotype for OPP13-CAPS, PR-CAPS, and CRF-SCAR, respectively. In contrast, we were able to detect male sterile cytoplasm in some breeding lines, and we were also able to detect polymorphisms for PR-CAPS between stable and unstable maintainer lines. The 17T7-SNP also showed polymorphisms between unstable and stable maintainer (or restorer) lines. In conclusion, we expect that it will be possible to develop stable A, B, and C sweet pepper lines using CMS-associated markers and that this will eventually lead to successful implementation of the CMS/Rf system to produce F1 hybrid sweet pepper seeds.

Isolation and Analysis of Rice Rf1-Orthologus PPR Genes Co-segregating with Rf3 in Maize

Using an in silico cloning approach, five putative maize pentatricopeptide repeat (PPR)-containing protein genes (PPR-814a, PPR-814b, PPR-814c, PPR-816, PPR-817) with complete open reading frames were identified in the inbred line S-Mo17Rf3Rf3. The amino acid sequence indicated that these genes encoded mitochondrially targeted proteins containing repeats of a 35-aa PPR motif. The genes were mapped into the interval umc1525–bnlg1520 on chromosome 2. In a non-restoring genotype, we identified three homologous genes that contained deletions or nucleotide substitutions in the coding region. Sequence analysis revealed that one of the three genes (PPR-814a, PPR-814b, PPR-814c) could be considered a candidate restorer gene for S male sterility cytoplasm, and linkage analysis demonstrated that the genes co-segregated with the fertility restorer gene Rf3.

Molecular Identification and Distinctness of NSa Male Sterile Cytoplasm in <I>Brassica napus</I>

Molecular Identification and Distinctness of NSa Male Sterile Cytoplasm in <I>Brassica napus</I>

Inheritance of Restorer Gene for CMS-FA Hybrid Rice

发掘水稻新型雄性不育细胞质源CMS-FA，育成系列优质米不育系和系列新质源恢复系，组配成强优势杂交稻组合的基础上研究新质源雄性不育恢复系的恢复基因遗传。采用新质源(CMS-FA)不育系金农1A与恢复系金恢3号杂交获得杂交F 1 代种子，种植F 1 代，收获自交F 2 代种子。用F 1 分别与不育系或保持系回交，获得(不育系//不育系/恢复系和不育系/恢复系//保持系)2个测交群体。同时种植P 1 、P 2 、F 1 、F 2 、B 1 F 1 和B 2 F 1 等群体，考察花粉染色率、套袋结实率和自然结实率，卡平方测验遗传分离适合度。结果表明，不育系与恢复系杂交F 1 代正常可育，育性恢复(可育)基因为显性遗传。F 2 代分离出可育︰不育适合3︰1，育性恢复(可育)基因为1对显性基因控制。B 1 F 1 和B 2 F 1 代2个测交群体的可育︰不育都适合1︰1分离规律，验证了F 2 代育性恢复(可育)单基因的遗传模式。暂时确定新质源(CMS-FA)核质互作三系的基因型为不育系S(SS)、保持系F(SS)和恢复系S(FF)。

Effect of sterile cytoplasm types on pigment content in leaves of F1 grain sorghum hybrids

Differences are found in the content of chlorophylls a and b and carotenoids in green leaves of F1 grain sorghum hybrids obtained on the basis of alloplasmic lines with genetically different types of CMS-inducing cytoplasms. Sterile cytoplasms A2, A4, and 9E cause a considerably higher content of chlorophylls in hybrids than cytoplasm A1 traditionally used in breeding sorghum.

Validation of SCAR markers, diversity analysis of male sterile (S-) cytoplasms and isolation of an alloplasmic S-cytoplasm in Capsicum

Abstracts Cytoplasmic male sterility (CMS) is increasingly being utilized for hybrid seed production of hot pepper ( Capsicum annuum ). Two CMS specific sequenced characterized amplified regions (SCARs), viz., atp6 607 and coxII 708 developed elsewhere were validated in an array of genotypes possessing male sterile (S-) and normal/male fertile (N-) cytoplasms and their feasible uses in CMS based pepper hybrid breeding have been elaborated. A set of eight maintainer and restorer inbreds were crossed on four CMS lines possessing two independently isolated and commercially utilized S-cytoplasms (Peterson's and Reddy's). Based on fertility restoration/maintenance reaction of 32 resulted F 1 s and on the presence of two SCARs ( atp6 607 and coxII 708 ) in both the S-cytoplasms, it has been concluded that although two S-cytoplasms were isolated and commercially utilized independently, they are genetically same or similar. Through inter-specific hybridization between C. chacoense and C. annuum , a new alloplasmic S-cytoplasm in the genus Capsicum has been isolated and the CMS F 1 has been advanced to CMS BC 1 F 1 .

Inter- and intraspecific sequence variations of the chloroplast genome in wild and cultivatedRaphanus

To assess the differentiation of the chloroplast genome in wild and cultivated species of Raphanus, nucleotide sequence polymorphisms were investigated for approximately 2 kbp ranging from trnL (UAA) to psbG in R. raphanistrum and R. sativus. Eighteen plants of wild species, 10 Japanese wild radish plants (R. sativus), and 31 cultivated plants were used for sequence analysis. Intraspecific variations of the chloroplast genome were present both in wild and cultivated Raphanus. All three genes investigated (trnL, trnF and ndhJ) contained nucleotide substitutions within the genus. Whereas, larger numbers of mutations were observed in the intergenic regions. Using the detected variations, the 59 radish plants were classified into 11 haplotypes, seven of which were unique to wild species. Among the haplotypes, one type corresponded completely with the Ogura male sterile cytoplasm. All the cultivated radishes belonged to one of four types, of which three were also observed in Japanese wild radish. The haplotypes were classified into four groups by cluster analysis, and the distribution in the dendrogram confirmed that cultivated radish has multiple origins. On the other hand, the seven haplotypes uniquely observed in R. raphanistrum were considered as useful materials to provide genetic diversity of cytoplasm for breeding of cultivated radishes.

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

Expression of the mitochondrial gene orf138 causes Ogura cytoplasmic male sterility (CMS) in Raphanus sativus, but little is known about the mechanism by which CMS takes place. A preliminary microarray experiment revealed that several nuclear genes concerned with flavonoid biosynthesis were inhibited in the male-sterile phenotype. In particular, a gene for one of the key enzymes for flavonoid biosynthesis, chalcone synthase (CHS), was strongly inhibited. A few reports have suggested that the inhibition of CHS causes nuclear-dependent male sterile expression; however, there do not appear to be any reports elucidating the effect of CHS on CMS expression. In this study, the expression patterns of the early genes in the flavonoid biosynthesis pathway, including CHS, were investigated in normal and male-sterile lines. In order to determine the aberrant stage for CMS expression, the characteristics of male-sterile anthers are observed using light and transmission electron microscopy for several stages of flower buds. The expression of CHS and the other flavonoid biosynthetic genes in the anthers were compared between normal and male-sterile types using real time RT-PCR. Among the flavonoid biosynthetic genes analysed, the expression of CHS was strongly inhibited in the later stages of anther development in sterility cytoplasm; accumulation of putative naringenin derivatives was also inhibited. These results show that flavonoids play an important role in the development of functional pollen, not only in nuclear-dependent male sterility, but also in CMS.

Qualitative and quantitative analysis of mitochondrial and cytoplasmic proteins

It has been established that the content of proteins in cytoplasm is 10 times higher than in mitochondria. Of the four analyzed genotypes, the male-sterile line is characterized by the minimum content of cytoplasmic and maximum content of mitochondrial proteins in all investigated organs. Treatment with gibberellins stimulates the biosynthesis of proteins in three genotypes and inhibits their biosynthesis in line RW637Rf, both in the cytoplasmic and mitochondrial fractions. By means of electrophoresis, a protein with a molecular mass of 16 kDa, related to cytoplasmic sterility in sunflower and being a product of mitochondrial gene orfH522, has been found in leaves and flower heads of male-sterile line SW501CMS and fertile line SW501 treated with gibberellins. Its content is higher in the mitochondrial fraction of these genotypes.

Linkage analysis between the partial restoration (pr) and the restorer-of-fertility (Rf) loci in pepper cytoplasmic male sterility

Cytoplasmic male sterility (CMS) in chili pepper is restored by one major dominant nuclear gene, restorer-of-fertility (Rf), together with some modifier genes and is also affected by temperature. As a result, male fertility was identified as having several phenotypes. That identified and used in the present study allowed partial restoration of fertility, producing plants that simultaneously produce normal and aborted pollen grains, with most grains stuck to the anther wall, even after dehiscence, resulting in low seed set per fruit. The trait was visible only in the presence of Paterson's sterile cytoplasm and was controlled by a recessive nuclear gene, partial restoration (pr). A CAPS marker, PR-CAPS, closely linked to the trait, has been developed by Lee et al. (2008). In this study, linkage analysis was performed in 205 F(2) individuals derived from the 'Buja' Korean commercial F(1) chili pepper variety using the PR-CAPS marker and the three Rf-linked markers (OPP13-CAPS, AFRF8-CAPS, and CRF-SCAR) previously reported. Consequently, we found that these four markers were tightly linked. This result means that the pr gene might be tightly linked to the Rf locus or the third allele of Rf locus. The sequence diversity of the pr- and Rf-linked markers was also analyzed. The internal sequences of OPP13-CAPS (1,180 bp) and PR-CAPS (640 bp) markers in 91 Korean inbred lines were clearly divided into three haplotypes. According to the sequencing results, a new PR-CAPS (MseI or SphI digestion) marker was designed to distinguish the three haplotypes. This marker will be useful for marker-assisted selection to develop new maintainers and restorers in commercial hybrid pepper breeding using CMS.

Unique High Molecular Weight Proteins expressed in Cytoplasmic Male Sterile Wheat (Triticum aestivum) as determined by Two‐Dimensional Electrophoresis (2DE)

When careful computer analysis of the patterns produced during 3 separate growth years by 2DE was performed the separation of expressed proteins at the young spike stage of growth of male sterile wheat plants indicated there were 18 proteins missing when compared to male fertile plants at the same growth stage. Surprisingly, there were found 12 high molecular weight (over 90 kDa) proteins present in the male sterile plant patterns that were not present in the male fertile plant patterns. Molecular weight and isoelectric points of the affected proteins will be presented. Leaf 2DE patterns at the corresponding developmental growth stages indicated there were no differences to be found. This suggested that the protein pattern differences seen are specifically expressed at the spike stage of growth. Searching of the literature suggests this is the first study of expressed proteins by 2DE of the rather common cytoplasmic male sterility phenomenon in nature. Because of this it is tentatively proposed that male cytoplasmic sterility in wheat at least may have a quite unique genetic program controlling expressed proteins in specific structures of wheat in at least part of its life cycle. Supported in part by institutional research grants from Harbin Normal University.

Analysis of Differential Expression of Mitochondrial Proteins between C-Type Cytoplasmic Male Sterility Line C48-2 and Maintainer in Maize

Cytoplasmic male sterility (CMS) in maize (Zea mays L.) is categorized into T-CMS, C-CMS, and S-CMS types based on the specific restoration of nuclear gene, in which C-CMS has a great application potential in maize hybrid breeding due to its stable sterility and resistance to Helminthosporium maydis Nisik. The sterile mechanism of C-CMS is not fully understood for its later discovery and utilization than these of T-CMS and S-CMS. The objective of the study was to give some evidences and explanations to the sterility reason of C-CMS in maize by analyzing the differential expression of anther mitochondrial proteome between a C-type sterile line (C48-2) and its maintainer (N48-2). The mitochondrial proteins in anthers of C48-2 and N48-2 were separated firstly by two-dimensional electrophoresis with immobilized pI (310, non-linear) gradients and then by SDS-PAGE. The coomassie brilliant blue-stained protein spots were analyzed using PDQUEST software, there were about 260 detectable spots on each 2D-gel. With direct MALDI-TOF mass spectrometry analysis and protein database searching, only 2 out of 10 protein spots were identified with significative data, which were up-regulated in mononuclear phase of C48-2. They were identified as glutamate dehydrogenase (GDH) and voltage-dependent anion channel protein-1 (VDAC1), respectively. The GDH exists mainly in mitochondria of higher plants and plays an important role in nitrogen metabolism. Once the GDH is up-regulated at any phase of plant development, the normal supply of energy will be affected, probably leading to cytoplasmic sterility. The VDAC1 plays an essential role not only in the permeability of the mitochondrial membrane, but also in apoptosis mediated by the mitochondrial pathway. Overexpression of VDAC1 in a variety of cells induced apoptotic cell death, indicating VDAC1 as a conserved mitochondrial element of the death machinery. Our research suggests that the GDH and VDAC1 in mitochondria probably cause the sterility in C-CMS in maize.