Identification and characterization of new sources of cytoplasmic male sterility in rice

Search for male sterility-inducing cytoplasm in wild species of the genus Oryza was attempted with a view to diversify the base of the cytoplasmic genetic male sterility system currently used in the development of commercial rice hybrids. A total of 132 interspecific crosses were made involving accessions of four wild and two cultivated species, all belonging to the A genome. Wild accessions possessing sterility-inducing cytoplasms were identified following reciprocal and sterile F₂ backcross methods. Sterile segregants were pursued through substitution backcrosses to develop cytoplasmic male-sterile (CMS) lines. CMS lines were developed with the cytoplasm of either O. rufipogon (VNI) or O. nivara (DRW 21018, DRW 21001, DRW 21039, DRW 21030 and RPW 21111). Based on shape, staining, and abortive pattern of pollen and also on type of interaction with a set of restorers and maintainers for known cytoplasmic male steriles of WA source (V 20A), the newly-developed CMS lines were grouped into four classes. Of these, RPMS 1 and RPMS 2 showed gametophytic male sterility with a restorer reaction different from WA cytoplasmic male-sterile stocks.     

Development of new cytoplasmic source of male sterile lines in rice for shallow lowland

Two shallow lowland male sterile lines, MotiA and PadminiAwith Miz.4 cytoplasmic source were developed throughintervarietal hybridization and recurrent backcrossing withtheir respective parents. In all probability, MotiA and PadminiA of Miz.4 source may be genetically different from the Wild abortive CMS source (V.20A). These two CMS lines are semitall in stature, photosensitive, of late duration in flowering with easy fertility restoration ability, and thus could be useful for developing hybrid rice suitable for shallow lowland areas.     

A new cytoplasmic male sterility source from wild Helianthus annuus

Summary A new cms source, ANN-5, was found in wild Helianthus annuus. This source showed high stability under different conditions in 1991 and 1992. All progenies from crosses of this source with several stable B-lines and restorer lines, which are homozygous for the gene which restores Leclercq's source of male sterility, were completely male sterile. Flower contained pistils and atrophied stamens. The cytological analysis showed that pollen mother cell degeneration took place in a premeiotic stage.     

Cytological observations in some hybrids between the rice species Oryza sativa L. and O. glaberrima Steud.

Summary Several hybrids between Oryza sativa and O. glaberrima and their backcrosses with O. sativa were studied. Their seed sterility was very different; large differences were also observed in the level of pollen sterility and in the earliness of microspore failure. The proportion of aborted embryo sacs was much lower than the rate of sterile male gametophytes. The backcross populations were much more sterile than the corresponding F₁ hybrids. On the base of our observations and according to the literature, we may conclude that genic unbalance is the main cause of sterility of these hybrids, but that physiological factors may also be involved. Thus a restoration of fertility is generally possible by selection. On the other hand, male-sterile lines could be bred from some of these hybrids.     

A new source of cytoplasmic male sterility in sunflower

Summary Interspecific substitutions of the nucleus of Helianthus annuus (2n=34) cv. Saturn into the cytoplasm of H. petiolaris (2n=34) by successive backcrossing, resulted in progenies with indehiscent anthers containing white, rather than normal yellow, pollen. Further backcrossing by cv. Saturn failed to restore pollen shed, suggesting that the male sterility was cytoplasmic. In vivo germination tests of pollen from 23 such plants from eight BC₅ lines, indicated complete pollen sterility for 14 plants, but normal seed set, suggesting that female fertility was not affected. Meiosis in all plants examined was normal.Crosses between seven sources of pollen-fertility restorer, one collection of wild H. annuus, and an existing source of cytoplasmic male sterility, resulted in a high frequency of plants with normal pollen shed in all F₁ progenies. However, no normal pollen shed was evident in F₁ progenies for similar crosses between BC₅ male-steriles and three of the seven restorer sources, nor for the single wild H. annuus evaluated. The foregoing suggests that the backcross substitution lines are a new source of cytoplasmic male sterility. The inheritance of restoration of pollen shed was complex and not fully elucidated. Some data suggested that two independent, complementary, dominant genes were required, but others indicated two to three independent, dominant genes.     

Genetics of thermosensitive genic male sterility in rice

Summary Inheritance of thermosensitive genic male sterility (TGMS) in Norin PL12 and IR32364TGMS and their allelic relationship were studied from F₁, F₂ testcross (TC) and F₃ generations of the crosses made with the two mutants and several fertile tester parents. F₂, TC and F₃ segregation behavior for pollen and spikelet fertility indicated that the TGMS trait in the two mutants was controlled by a single recessive gene. Allelic relationship studies indicated that TGMS genes of the two mutants were different. Since TGMS gene in Norin PL12 has been designated as tms ₂ , the TGMS gene present in IR32364TGMS is tentatively designated as tms ₃ (t) until allelic test is done with another TGMS gene (tms ₁ ) reported from China in a line 5460S seeds of which were not available.     

Interspecific cytoplasm substitutions of an Indica strain of Oryza sativa L. and O. Glaberrima Steud

Summary To study differential nucleus-cytoplasm interactions between the two cultivated rice species, Oryza sativa and O. glaberrima, cytoplasmic substitution lines were made by using a glaberrima strain (G) and an Indica strain of sativa (S). The G cytoplasm had no adverse effect on pollen development when combined with the nucleus of S. On the other hand, when the S cytoplasm was combined with the G nucleus, the substitution line showed no seed set because of male sterility although the pollen grains were normally stained with I₂-KI solution. A dominant gene derived from S strain seemed to cause anther indehiscence in the substitution line. Further, a restorer gene (Rf _j)from Akebono of Japonica type was effective on pollen restoration in the male sterile line, suggesting that the S cytoplasm is the same as those of Japonica type in terms of a fertility-restoring system.This paper is Genetic studies of speciation in cultivated rice. 4.     

Cytoplasmically controlled cold tolerance in a cytoplasmic-genetic male sterile line of rice

Summary The cytoplasmic-genetic male sterile line, Krishna-A with Kalinga-I cytoplasm was developed in rice through repeated backcrossing the completely pollen sterile F₁ of the cross Kalinga-I/Krishna with the recurrent male parent cv. Krishna. The germination percentage and root-shoot length in Krishna-A at 12°C and 9°C indicated a higher degree of tolerance to low temperature than the cold tolerant female parent Kalinga-I. The high yielding cold tolerant variety Kalinga-I was developed from a cross involving the cold tolerant variety Dunghansali and the high yielding variety IR 8. It is inferred that tolerance to low temperature during germination in Krishna-A was inherited from the cytoplasm of cv. Dunghansali through Kalinga-I besides male sterility. The cytoplasmic control of cold tolerance is reported here for the first time in rice literature. The male sterile line Krishna-A because of its tolerance to low temperature would be suitable for the development of hybrid rice especially for areas where low temperature is a problem during germination.     

Two linked genes on rice chromosome 2 for F1 pollen sterility in a hybrid between Oryza sativa and O. glumaepatula

Hybrid incompatibility plays an important role in establishment of post-zygotic reproductive isolation. To unveil genetic basis of hybrid incompatibilities between diverged species of genus Oryza AA genome species, we conducted genetic dissection of hybrid sterility loci, S22(t), which had been identified in backcross progeny derived from Oryza sativa ssp. japonica (recurrent parent) and South American wild rice O. glumaepatula near the end of the short arm of chromosome 2. The S22(t) region was found to be composed of two loci, designated S22A and S22B, that independently induce F₁ pollen sterility. Pollen grains containing either of the sterile alleles (S22A-glum^s or S22B-glum^s) were sterile if produced on a heterozygous plant. No transmission of the S22A-glum^s allele via pollen was observed, whereas a low frequency of transmission of S22B-glum^s was observed. Cytological analysis showed that the sterile pollen grains caused by S22A could reach the bicellular or tricellular stage, and the nearly-sterile pollen grains caused by S22B could reach the tricellular stage. Our genetic analysis showed repulsion linkage effect is possible to induce strong reproductive barrier by high pollen sterility based on recombination value and transmission ratio of hybrid sterility gene to the progeny was influenced by frequency of competitors on fertilization.     

Chromosomal location of fertility restoring genes for ‘wild abortive’ cytoplasmic male sterility using primary trisomics in rice

Summary Identification and location of fertility restoring genes facilitates their deployment in a hybrid breeding program involving cytoplasmic male sterility (CMS) system. The study aimed to locate fertility restorer genes of CMSWA system on specific chromosomes of rice using primary trisomics of IR36 (restorer), CMS (IR58025A) and maintainer (IR58025B) lines. Primary trisomic series (Triplo 1 to 12) was crossed as maternal parent with the maintainer line IR58025B. The selected trisomic and disomic F₁ plants were testcrossed as male parents with the CMS line IR58025A. Plants in testcross families derived from disomic F₁ plants (Group I crosses) were all diploid; however, in the testcross families derived from trisomic F₁ plants (Group II crosses), some trisomic plants were observed. Diploid plants in all testcross families were analyzed for pollen fertility using 1% IKI stain. All testeross families from Group I crosses segregated in the ratio of 2 fertile: 1 partially fertile+partially sterile: 1 sterile plants indicating that fertility restoration was controlled by two independent dominant genes: one of the genes was stronger than the other. Testcross families from Group II crosses segregated in 2 fertile: 1 partially fertile+ partially sterile: 1 sterile plants in crosses involving Triplo 1, 4, 5, 6, 8, 9, 11 and 12, but families involving triplo 7 and triplo 10 showed significantly higher X² values, indicating that the two fertility restorer genes were located on chromosome 7 and 10. Stronger restorer gene (Rf-WA-1) was located on chromosome 7 and weaker restorer gene (Rf-WA-2) was located on chromosome 10. These findings should facilitate tagging of these genes with molecular markers with the ultimate aim to practice marker-aided selection for fertility restoration ability.     

Isolation and genetic characterization of a fertility-restoring revertant induced from cytoplasmic male sterile rice

Summary A male fertile revertant was isolated from M₁ of a cytoplasmic male sterile indica rice line II-32A, the dry seeds of which were treated with ⁶⁰Co- rays at a dose of 290 Gy. The acquired revertant T24 was morphologically and agronomically similar to II-32B, the maintainer of II-32A. Testcrosses of the revertant with II-32A and Zhenshan 97A showed that the revertant was able to restore the fertility of CMS lines. Genetic analysis of the progenies between T24 and II-32A, Zhenshan 97A XieqingzaoA and DZhenshan 97A, which have different cytoplasms, showed that the fertility restoration of four CMS lines by T24 involved one nuclear gene, indicating that T24 was a result of the mutation of one nuclear gene. The mechanism of the restoration of CMS line by T24 was obviously different from other restorers such as Minghui 63 and 20964, which were shown to behave in two gene mode in fertility restoration. The discovery of the revertant T24 contributes to the understanding of CMS and fertility restoration of CMS in rice. The T24 and its parent II-32A may constitute a pair of near isogenic lines for the restoring gene, which should be valuable materials for molecular genetic analysis of CMS.     

Increasing biodiversity of irrigated rice in Africa by interspecific crossing of Oryza glaberrima (Steud.) × O. sativa indica (L.)

The African rice Oryza glaberrima, traditionally cultivated since more than 3.500 years, is of poor agronomic performance but resistant/tolerant to various stresses and diseases. The introduction of these characters into O. sativa cultivars is difficult since crossing barriers cause spikelet sterility in F1. Backcrossing can restore fertility and recently facilitated the development of fertile O. glaberrima × O. sativa ssp. japonica hybrid progenies for rain fed systems. With the objective to gain access to African rice germplasm for improvement of irrigated rice, crosses were performed with eighteen O. glaberrima and twenty O. sativa ssp. indica accessions. In total about one hundred F1-hybrid grains were obtained. The F1 plants were all completely sterile and backcrossing (BC) to O. sativa was performed in order to restore spikelet fertility. Monitoring of Tog5681 × IR64 hybrid progenies under field conditions revealed a broad genetic diversity within the BC₁ and BC₂ populations. Some BC₁ and BC₂ progeny plants outperformed the O. sativa parent, indicating that the heterozygocity level and complementary gene action after two backcrosses are still sufficient to positively influence plant vigor. Spikelet fertility of progenies was highly variable, but almost complete fertility was already observed within the BC₁F₂ population. High spikelet fertility was preserved in one out of two analyzed BC₁F₃ families and inmost of the BC₂F₃ families. The ability to restore spikelet fertility within few generations and the potential of the genetic diversity present in interspecific progenies facilitates the development of plant types specifically designed for the African irrigated and lowland environment. This revised version was published online in August 2006 with corrections to the Cover Date.     

Development of new cytoplasmic genetic male sterile lines through Indica × Japonica hybridization in rice

Summary From 28 Indica-Japonica crosses, two Indica cultivars, V.20B and Sattari were identified to possess male sterile cytoplasm with fertility restoring genes. It was possible to develop a new Japonica cytoplasmic genetic male sterile line (Zhunghua-1) on Indica male sterile cytoplasm (V 20B) by repeated backcrossing the complete pollen sterile plants of V 20B x Zhunghua-1 to the recurring male parent, Zhunghua-1. The study indicated that it would be possible to develop male sterile lines rom indica-japonica crosses only when there is sufficient amount of reciprocal differences with respect to pollen sterility. Further, it was inferred that it would be easier to develop Japonica male sterile lines on Indica cytoplasm than developing Indica male sterile line with japonica cytoplasm.     

Cytoplasmic-genetic male sterility in radish (Raphanus sativus L.). Identification of maintainers,inheritance of male sterility and effect of environmental factors

Summary Research has been carried out on identification of maintainers for cytoplasmic-genetic male sterile lines in Japanese and European radish, on the mode of inheritance of male sterility and on the effect of environmental factors on the expression of this character.In a Japanese radish population and in most early European radish populations maintainers were found in high frequency. Segregations for male sterility in full-sib families, obtained by crossing male sterile and male fertile plants, and in backcross generations, indicated that male sterility is probably determined by one dominant and two recessive independently acting genes, but also minor genes may be involved.The expression of male sterility was not affected by seasonal influences. In some populations a reversible temperature effect was found, most ms plants occurred at 10, 14 and 26°C and most mf plants at 17 and 20°C.     

A novel environment-induced genic male sterile (EGMS) mutant in indica rice

Summary Male-sterile mutants were isolated from M₂ and M₃ generations of indica rice variety 26 Zhaizao, dry seeds of which had been exposed to ⁶⁰Co- rays at a dose of 290 Gy. The mutants were planted in early season and ratooned in late season for two successive years for identification of fertility conversion in different growing seasons. One of the mutant lines was further observed in a growth chamber and in the field. Results showed that daily average temperature might be the major factor conditioning the male fertility conversion at a moderate daylength. The critical temperature for the male fertility conversion of the mutant grown under 12.5 h and 14.0 h daylength is about 23°C, below which the plant becomes completely male sterile. Its male fertility conversion character differs from other EGMS lines so far developed. The performance of the hybrids between the mutant and some other indica varieties demonstrated its good combining ability and its potential value in hybrid rice production. The obtained mutant line still sheds KI-stainable pollen grains under male sterilizing conditions. Nevertheless, pollen grains shed from the male sterile plants were much more vulnerable than from normal plants. At sucrose concentration below 1.5 M, the pollen grains from the mutant grown under male sterilizing conditions almost completely broke down, while above 1.5 M they became plasmolysed and shrunken. This is indicative of poor development of the membrane and walls of the pollen grains from the male sterile mutant, causing the pollen grains to be unfunctional. NBT test also clarified the abortion of the pollen grains from the mutant, which were formed in the male sterilizing environment.     

Wide compatibility in rice (Oryza sativa L.)

Summary Wide compatible varieties (WCVs) show normal spikelet fertility in crosses with Indica and Japonica rice varieties. Crosses of Indica and Japonica varieties frequently show high spikelet sterility which prevents exploitation of heterosis for grain yield. We screened 41 rice varieties for the wide compatibility trait by crossing each with three Indica and three Japonica testers. Varieties giving fertile F₁ hybrids with both groups of testers were classified as WCVs. Seven varieties viz., BPI-76 (Indica); N 22; Lambayeque-1 and Dular (Aus); Moroberekan, Palawan and Fossa HV (Japonicas), were identified as WCVs. The frequency of WCVs was higher among Aus and Japonicas. The wide compatibility trait in varieties: Dular and Moroberekan was controlled by a single dominant gene linked with the Est-2 and Amp-3 loci (mean recombination 32.0%). Est-2 and Amp-3 showed complete linkage. Pgi-2 was found to be linked with Est-2 and Amp-3 (mean recombination 16.1%). Est-2 and Amp-3, showed a tighter linkage with C ⁺ (mean recombination 4.1%). Pgi-2 showed a lower linkage with C ⁺ (mean recombination 17.3%). The recombination values between the WC gene in Dular and C ⁺ was much higher than those reported in Japan for the WC gene (S₅ ⁿ) from Ketan Nangka. It is possible that the WC gene from Dular is different from that in Ketan Nangka. Linkage intensities with the WC gene were not strong enough to be of use for indirect selection for the wide compatibility trait. A search for a more closely linked isozyme or DNA marker was proposed.     

Identification of two types of differentiation in cultivated rice (Oryza sativa L.) detected by polymorphism of isozymes and hybrid sterility

Magnitudes of genetic variation within each of major varietal groups of cultivated rice were surveyed in terms of isozyme polymorphism and allelic differentiation of hybrid sterility loci, both of which are considered to have litt le selective value. Allelic differences for 20 isozyme loci were examined in a total of 337 accessions, including Indica and Japonica rices. Aus cultivars from India, Javanica cultivars and both landraces and cultivars from China. Eleven out of the 20 isozyme loci were polymorphic. The Aus cultivars contained more alleles per locus than others. The hybrid sterilities in the crosses among Chinese cultivars, Indica and Japonica cultivars were mainly controlled by locus S-5, whereas the hybrid sterilities of Aus cultivars, when they were crossed to Indica, Japonica or Javanica cultivars, were found to be controlled by allelic interaction at hybrid sterility loci S-5, S-7, S-9 and S-15. Also in terms of the number of alleles at S-5 and S-7, Aus cultivars contained more alleles than others. While the Aus group showed an extreme diversity for both hybrid sterility alleles and isozymes, modern cultivars from the south of YangZi River in China were classified into Indica type and those from the north were into Japonica, which were almost the same as those in Japan. On the basis of the measured polymorphism the Indica-Japonica differentiation was explained by founder effects, i.e., through selection and distribution of two original sources each with a unique set of markers. Contrastingly, the continuous and pronounced diversity in the Aus group was attributed to the absence of such a process. The intermediate groups in Yunnan province and Tai-hu Lake region of China are considered to be isolated from such founder effects, retaining an intermediate diversity in terms of isozymes and hybrid sterility genes. Since hybrid sterility hampers the exchange of genes between cultivars of different groups, the understanding of its genetic basis will be important in rice breeding, particularly in hybrid rice breeding. This revised version was published online in July 2006 with corrections to the Cover Date.     

Development of new sources of cytoplasmic male-sterile lines in rice

Two male-sterile lines, KalashreeA and PadminiA, with a Miz.21 cytoplasm source were developed through indica/indica hybridization followed by repeated backcrossing with their respective recurrent male parents (Kalashree and Padmini) up to the BC₆ generation. These two cytoplasmic male-sterile lines are suitable for use in the development of hybrids for lowland situations owing to their intermediate to semi-tall stature, late flowering duration, good grain quality and easy fertility restoration ability.     

Photoperiod-sensitive cytoplasmic male sterility in wheat with Aegilops crassa cytoplasm

Summary Triticum aestivum cv. Norin 26 with Aegilops crassa, Ae. juvenalis or Ae. vavilovii cytoplasm (all D² type) has been studied relative to its photoperiodic response of male sterility and fertility restoration patterns. Alloplasmic lines of Norin 26 with a D² type cytoplasm showed almost complete male sterility under long-day conditions (15 h), but high male fertility under short-day conditions (14.5 h). No significant influence of temperature on reduction in male fertility was observed. Thus, this type of male sterility is called photoperiod-sensitive cytoplasmic male sterility (PCMS). The PCMS is expressed in the form of pistillody of stamens. Histological studies revealed that there were incomplete ovule-like structures instead of tapetal cells and pollen grains in the pistillate stamens. The floret differentiation stage of the plant is the stage that is sensitive to photoperiod. The PCMS can be used as a new means for hybrid wheat production, named two-line system. The PCMS line is maintained and multiplied by self-fertilization under short-day conditions, and hybrid seed can be produced by crossing the PCMS line with a pollinator line under long-day conditions. In contrast to the system of hybrid wheat production using the T. timopheevi cytoplasm, the present system requires only PCMS and pollinator lines.Abbreviation CS Chinese Spring - N26 Norin 26 - PCMS photoperiod-sensitive cytoplasmic male sterility