Timopheevi Cytoplasm Research Articles

Usefulness of alien sterilizing cytoplasms for the hybrid breeding of triticale (xTriticosecale Wittmack): preliminary results.

To be useful for cereal breeding, cytoplasmic male sterility (CMS) should express the complete sterility of maternal lines and the full restoration of the male fertility of F1 hybrids. The most reliable source of sterilizing cytoplasm for triticale is Triticum timopheevi; however, due to the low frequency of efficient non-restorer genotypes for this cytoplasm, new sources of CMS are needed. In this study, aside from T. timopheevi (T) cytoplasm, three alternative CMS sources were tested: Pampa (P) from Secale cereale L., Aegilops sharonensis (A), and Ae. ventricosa (V). The suitability of these cytoplasms for breeding was assessed based on the male fertility/sterility of F1 hybrids obtained through the manual pollination of CMS maternal lines with 36 triticale cultivars and breeding strains. About half of the hybrids with each type of cytoplasm were fully fertile and produced more than 30 grains per bagged spike. The highest percentage was found in hybrids with P cytoplasm (58.33%) and the lowest in hybrids with A cytoplasm (44.44%). Male sterility was observed in hybrids with P cytoplasm (16.67%) and A cytoplasm (16.67%) but not in hybrids with T or V cytoplasm. In terms of practical aspects, male sterility systems with P or A cytoplasm exhibit similarity in their ability to restore male fertility that differ from the T and V cytoplasms. Although all studied cytoplasms exhibited some disadvantages for breeding purposes, none should be definitively classified as unacceptable for future breeding programs regarding the development of triticale hybrid cultivars.

Male sterility of triticale lines generated through recombination of triticale and rye maintainers

The Triticum timopheevi cytoplasmic male sterility (cms) system in triticale (xTriticosecale Wittmack) suffers from a low frequency of maintainers and environmental instability of the male sterility. On the other hand, the Pampa cms system in rye (Secale cereale) exhibits strong male sterility and a low frequency of restorers. Here, we report generating hybrids between maintainers of the T. timopheevi cms system in triticale and maintainers of the rye Pampa cms system. Ten hybrids were obtained. Their hybridity was verified by PCR (polymerase chain reaction) using ISSR (inter simple sequence repeats) primers. The cms maintaining ability of F2 individuals and their progeny was tested. The F2 plants were crossed to male sterile lines of triticale carrying the T. timopheevi cytoplasm. Among 180 G1 offspring of these crosses, 71 (39.4%) were completely male sterile. Fourteen F2 individuals (7.8%), as well as their F2S1 and progeny, generated stable male sterility in G1, G1BC1 and G1BC2 generations after the crosses. Our results suggest that it is possible to produce a more stable cms system in triticale based on the T. timopheevi cytoplasm as compared to the existing one.

Genetic analysis and molecular mapping of a new fertility restorer gene Rf8 for Triticum timopheevi cytoplasm in wheat (Triticum aestivum L.) using SSR markers

A study on mode of inheritance and mapping of fertility restorer (Rf) gene(s) using simple sequence repeat (SSR) markers was conducted in a cross of male sterile line 2041A having Triticum timopheevi cytoplasm and a restorer line PWR4099 of common wheat (Triticum aestivum L.). The F1 hybrid was completely fertile indicating that fertility restoration is a dominant trait. Based on the pollen fertility and seed set of bagged spikes in F2 generation, the individual plants were classified into fertile and sterile groups. Out of 120 F2 plants, 97 were fertile and 23 sterile (based on pollen fertility) while 98 plants set ≥ 5 seeds/spike and 22 produced ≤ 4 or no seed. The observed frequency fits well into Mendelian ratio of 3 fertile: 1 sterile with χ(2) value of 2.84 for pollen fertility and 2.17 for seed setting indicating that the fertility restoration is governed by a single dominant gene in PWR4099. The three linked SSR markers, Xwmc503, Xgwm296 and Xwmc112 located on the chromosome 2DS were placed at a distance of 3.3, 5.8 and 6.7 cM, respectively, from the Rf gene. Since, no known Rf gene is located on the chromosome arm 2DS, the Rf gene in PWR4099 is a new gene and proposed as Rf8. The closest SSR marker, Xwmc503, linked to the Rf8 was validated in a set of Rf, maintainer and cytoplasmic male sterile lines. The closely linked SSR marker Xwmc503 may be used in marker-assisted backcross breeding facilitating the transfer of fertility restoration gene Rf8 into elite backgrounds with ease.

QTL analysis of fertility-restoration against cytoplasmic male sterility in wheat.

The cytoplasm of Triticum timopheevi causes cytoplasmic male sterility (CMS) in common wheat (T. aestivum) cv. 'Chinese Spring' (CS), and that of Aegilops kotschyi causes CMS in spelt wheat (T. spelta) var. duhamelianum (Sp). CS has fertility-restoring (Rf) genes against the latter cytoplasm and Sp has the ones against the former. To know the genetic system concerning to CMS, we crossed 66 F8 recombinant inbred lines (RILs) derived from a cross between CS and Sp as males to the alloplasmic lines of CS and Sp having the cytoplasms of T. timopheevi and Ae. kotschyi, respectively. The fertilities of respective F1 plants derived from the crosses were examined for QTL analysis. The major QTLs detected in both systems were located on the short arm of chromosome 1B. One minor QTL on chromosome 2B was also commonly detected in both of the systems, while other minor QTLs against T timopheevi cytoplasm were distributed on the chromosomes 2A, 4B, and 6A.

High-resolution RFLP mapping of the fertility restoration (Rf3) gene against Triticum timopheevi cytoplasm located on chromosome 1BS of common wheat.

Near-isogenic lines of alloplasmic wheat for the Rf3 gene controlling fertility restoration against the cytoplasm of Triticum timopheevi were developed by successive backcrossing of Chinese Spring to the F1 plants between (timopheevi)-CS (male sterile) and T. spelta (carrying Rf3). The resultant three BC3F1 plants were self-pollinated so as to obtain 125 BC3F2 progenies. Using this population, we precisely mapped the Rf3 gene on the short arm of wheat chromosome 1B (1BS) of the RFLP linkage map. The Rf3 gene was localized at a position 1.2 cM and 2.6 cM distant from Xcdo388 and Xabc156, respectively. The genetic distances of Rf3 from Nor and Gli-B1 were calculated to be 22.3 cM and 18.6 cM, respectively, supporting previous data. Estimation of the physical distance of the region suggets that the Rf3 gene resides within 500 kbp from the adjacent RFLP markers. The marker order of the genetic map corresponded to that of the cytological map, except for the position of Xbcd98. Comparison between genetic and cytological maps clearly shows that RFLP markers are unevenly distributed throughout the chromosome arm, and recombinations took place unequally in the chromosome arm.

Mitochondrial DMA variation in plants regenerated from embryogenic callus cultures of CMS triticale

The mitochondrial DNA (mtDNA) organization of primary hexaploid cytoplasmic male-sterile (CMS) triticale regenerants containing Triticum timopheevi cytoplasm was analysed by hybridization experiments and compared with the mitochondrial genome organization of the corresponding regenerants with maintainer cytoplasm. Callus cultures had been derived from immature embryos, and 623 triticale plants were regenerated via somatic embryogenesis after three to four subcultures. The chondriome of 159 regenerants was investigated with regard to somaclonal variation. Six different mitochondrial gene probes and four different restriction enzymes were used for Southern blot analyses by the non-radioactive digoxigenin labeling technique. Alloplasmic regenerants showed a gain or loss of hybridization signals up to a high percentage, while euplasmic ones revealed only minor variability with respect to band stoichiometries. In 24 cases rearrangements in the mtDNA were proved. We suppose that recombination processes and selective amplification events are responsible for these findings.

Non-radioactive organization and transcript analysis of the ATPase subunit 6 gene region in the mitochondrial genome from fertile and sterile (CMS) forms of wheat and triticale

In sterile triticale forms [with cytoplasmic male-sterile (CMS)-inducing timopheevi cytoplasm], fertile orms (with normal cytoplasm) and the corresponding wheat cross parents mitochondrial DNA (mtDNA) and RNA (messenger mtRNA) were characterized using total DNA and RNA material for Southern and Northern blots. A novel non-radioactive technique was applied by marking the probes with digoxenin. The fertile and sterile Triticum and triticale forms were analysed in three genes, atp 6, coxIII and rps13. These forms can be distinguished in the apt6 gene at the mtDNA and mtRNA levels.

Localization of Genes in Rye that Restore Male Fertility to Hexaploid Wheat with timopheevi Cytoplasm

AbstractFour sets of wheat‐rye addition lines were screened to localize genes in rye that restore male fertility to hexaploid wheat with timopheevi cytoplasm. One gene, designated Rfc3, was physically located in the distal 40 % of the long arm of chromosome 6R. No allelic variation at Rfc3 was found; normal male fertility was consistently observed in all F1 hybrid combinations tested. A second gene, designated Rfc4, was located on the long arm of chromosome 4R. Variation between chromosomes 4R in the level of restoration was observed; fertility in hybrids ranged from 0 % to about 50 % of normal. Attempts to genetically map Rfc4 were inconclusive but suggested it was located 16.1 cM from the telomere of the long arm and at least 8.0 cM from the centromere. These restorers, particularly Rfc3, may have potential in hybrid wheat breeding programs and can be manipulated for production of male sterile triticale lines.

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

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.

Incorporation of restoring gene of Aegilops umbellulata into wheat

Six 'Chinese Spring' – Aegilops umbellulata Zhuk. addition lines (UAD, UBD, UCD, UDD, UED, and UFM) were assayed for their effects on the fertility of timopheevi cytoplasm male sterile lines (T-type). Chromosome 6U of disomic addition line UAD was found to be able to restore the fertility of T-type male sterility and 'Chinese Spring' was verified to lack restoring genes, indicating that 6U carries at least one fertility restoration gene. From about 200 plants with 42 somatic chromosomes derived from the progeny of crosses Qu Xian Early A × UAD and Sumai No. 3 A × UAD, eight self-fertile plants were selected. Their self-fertility in timopheevi cytoplasm implies that they carry the restoring gene(s) from 6U. Cytological analysis was conducted on the hybrid F1 of the selected fertile plants (040-5, 060-1, and 061-4) as female parents crossed with 'Chinese Spring'. The self-fertility segregation and the chromosome pairing of pollen mother cells of F1 fertile plants from 040-5, 060-1, and 061-4 × 'Chinese Spring' during meiosis suggested that they were heterozygous translocation lines with restoring gene(s) from 6U.Key words: Aegilops umbellulata Zhuk., restoring genes, alien gene transfer, timopheevi cytoplasmic male sterile fertility, Triticum aestivum L.

The effect of the Triticum timopheevi nucleo-cytoplasmic system on the performance of an F1 hybrid wheat

Hybrid performance may be affected by the T. timopheevi nucleo-cytoplasmic system for producing hybrid wheat. Its effects were investigated in three experiments which compared the same hybrid genotype produced either in T. aestivum cytoplasm or in T. timopheevi cytoplasm. Few consistent differences were found between the two hybrids. The only significant difference in grain yield was associated with a significantly reduced seed set in the T. timopheevi hybrid, and suggested a likely effect of location on the degree of fertility restoration achieved. The T. timopheevi nucleo-cytoplasmic system appeared to reduce plant stand and test weight, but to improve flour colour and flour protein content relative to the T. aestivum hybrid.

Diallel analysis of androgenetic plant production in hexaploid Triticale (X. triticosecale, Wittmack)

Studies were made on the genetic determination of androgenetic plant yield and its two components: embryo production and green plant regeneration. This involved the analysis of a complete 7×7 diallel cross of 4 androgenetic lines and 3 lines obtained by pedigree selection, one of them having the Triticum timopheevi cytoplasm. The three traits analysed are both heritable and environmentally influenced (by season and culture medium composition). The analysis of embryo production shows a mainly nuclear inheritance, with predominantly additive gene action, but also a favourable effect of Triticum timopheevi cytoplasm. Green plant regeneration has a more complex genetic determination, with additive as well as non-additive gene action and cytoplasmic influences. Hybrids appear superior to inbred lines for embryogenesis and green plant yield, but not for green plant regeneration. Androgenetic lines used as parents did not show superiority over other parents either in their own value or in the transmission of androgenetic abilities. Genetic improvement seems to be possible by recombination in progenies of hybrids between lines having complementary abilities.

Agronomic and Male‐Fertility Restoration Characteristics of Wheat Restorer Lines with Aegilops speltoides and Triticum timopheevi Cytoplasms1

The use of Aegilops speltoides Tausch cytoplasm in fertility restorer line breeding was evaluated by comparing the male fertility restoration and agronomic performance of segregating population pairs in Ae. speltoides and Triticum timopheevi Zhuk. cytoplasms. Each of 15 restorer/restorer (R/R), eight restorer/cultivar (R/B), and two male‐sterile/restorer (A/R) F1's was crossed onto ms(timopheevi)‘Butte’ and ms(speltoides)Butte or ms(timopheevi)ND537 sib and ms(speltoides)ND537 sib to obtain populations with comparable nuclear gene segregation in the two cytoplasms. There was no significant difference between F1 seed set of populations with T. timopheevi cytoplasm and those with Ae. speltoides cytoplasm (82% vs. 76%). F3 bulks from fertile plants in each population were evaluated in replicated yield trials. Populations in the two cytoplasms performed similarly for all agronomic and seed quality traits except anther extrusion. Populations with T. timopheevi cytoplasm had 10% higher mean anther extrusion than populations with Ae. speltoides cytoplasm. We believe Ae. speltoides cytoplasm can be used interchangeably with T. timopheevi cytoplasm in hybrid wheat breeding and provides an alternative which can effectively broaden cytoplasmic variability.

Investigation of possible associations between partial fertility restoration and agronomic traits in Triticum aestivum L.

Many conventional hard red spring wheat (Triticum aestivum L. em Thell) lines, including several North Dakota cultivars, carry a gene (or genes) which restore partial male fertility to male sterile plants with Triticum timopheeviZhuk. cytoplasm. Since this gene has no fertility restoration function in T. aestivum cytoplasm, the postulation can be made that it is being retained in conventional lines because of pleiotropic effects, favorable linkages or chance. The research reported in this paper examined these possibilities. Forty F6 lines, derived from a single F2 plant which was heterozygous for a gene (or genes) for partial fertility restoration, were evaluated for two years in a yield trial planted at Fargo, North Dakota. The 40 lines were testcrossed to a male sterile line having T. timopheevi cytoplasm, and the mean seed set of testcrosses was used as a measure of a line's fertility restoration potential. Twenty-seven lines had the gene for partial fertility, and 13 lines apparently lacked this gene. The 40 lines differed for heading date, anther extrusion, plant height, grain yield, 200-kernel weight, test weight, and grain protein percentage. However, comparisons of lines having the restorer gene with those lacking the gene did not provide any obvious explanation for the retention of the partial fertility restorer gene in the breeding stocks of the North Dakota conventional hard red spring wheat breeding program. The possibility that the restorer gene was linked with genes for resistance to stem rust or leaf rust also was evaluated by testing lines for their reaction to several races of rust. No conclusive association was found.

Anther Ontogeny in an Exotic and Six Indigenous Cytoplasmic Male Sterile Lines of Wheat (Triticum aestivum) Possessing Triticum timopheevi Cytoplasm

Anther ontogeny in an exotic cytoplasmic male-sterile (CMS) Iine 'Nadadores' and that in six indigenous CMS Iines obtained through crosses with the former have been compared. In the exotic CMS Iine, pollen abortion was associated with nondegenerating and persistant tapetum. Similar abnormalities were recorded in the indigenous CMS Iines also. Besides a few other tapetal abnormalities like degeneration of tapetum in premeiotic stages, tapetal hypertrophy in both pre and post-meiotic stages and pseudo-periplasmodium formation were also recorded. It is concluded on the basis of these observations that malfunctioning tapeta of various types are caused by inhibition of vascular tissue of the anther connective.

Basic studies on hybrid wheat breeding

The nuclei of 12 common wheats (genome constitution AABBDD) were placed into the cytoplasms of Aegilops kotschyi and Ae. variabilis (both C(u)C(u)S(v)S(v)) by repeated backcrosses. Using these nucleus-cytoplasm hybrids, male sterility-fertility restoration relationship was investigated. Male sterility was expressed by these cytoplasms only in Slm, Splt and Mch. The other nine common wheat nuclei gave normal fertility against these cytoplasms. These cytoplasms were compared with the Triticum timopheevi cytoplasm that is now widely used in the hybrid wheat breeding program in order to investigate their effects on important agronomic traits of the 12 common wheats: The kotschyi and variabilis cytoplasms were as good as the timopheevi cytoplasm in this respect.The F1 hybrid between (kotschyi)- or (variabilis)-Splt and CS showed normal fertility. Segregation of the fertiles and steriles in their F2 generations followed the simple Mendelian fashion, i.e., 3 fertile∶1 sterile. Thus, the fertility restoration in this case is mainly controlled by a single dominant gene which will be designated as Rfv1. To determine its location, ditelo-lBS and -lBL of CS were crossed as male parents to male sterile (kotschyi)- and (variabilis)-Splt. The F1 hybrids between the male sterile Spit's and CS ditelo-lBS became male fertile, while those between the male sterile Spit's and CS ditelo-lBL became completely male sterile. Thus, the location of the gene Rfv1 has been determined to be on the short arm of chromosome lB of CS. Furthermore, a close relationship between the fertility-restoring genes and the nucleolus organizer region was pointed out.Finally, the schemes of breeding the male sterile lines of a cultivar with these cytoplasms, and its maintainer line were formulated. The following two points were considered as the advantages of the present male sterility-fertility restoration system over that using the timopheevi cytoplasm in breeding hybrid wheat: (1) easier fertility restoration in F1 hybrids, and (2) no need of breeding the restorer line.

Evaluation of three sources of pollen fertility restoration genes and natural outcrossing of cytoplasmic male-sterile wheat

Fertility restoration genes in Triticum aestivum L. in Texas Restorer Composite (TRC), D6301, and four CIMMYT restorer lines were studied, and selection was made for higher fertility in TRC. Mean-while, outcrossing percentages of seed set for 27 spring habit cytoplasmic male sterile (cms) varieties were evaluated for 3 to 5 years at Davis. The winter-habit TRC material did not restore reasonably good fertility, and the response to selection for higher fertility seemed to be slow. This poor fertility could be partly due to its late winter growth habit causing flowering at a period of high temperature and low humidity at Davis. The highest F1 fertility was 46.6% in the cross cms Ramona x TRC-6, and its F2 segregated into the ratio of 15 fertile to 1 sterile, with fertility ranging from 3.2 to 100%. Suggested for its improvement was intensive selection in the original TRC material and in the segre-gating F2 population, followed by intercrossing. D6301 has 2 fertility restoration genes with different strengths which restore fertility up to 45.2% when both genes are heterozygous. D6301 is quite likely heterogeneous for these genes. Four CIMMYT restorer lines, D7464, D7465, D7466, and D7467, had satisfactory F1 fertility restoration after crossing with cms Ramona 50. In 1975, the fertilities of the F1's ranged from 71 to 85% and were over 90% in 1976. The F2 population of the cross cms Ramona 50 × D7464 segregated into a ratio of 3 fertile to 1 sterile, indicating that D7464 has a single dominant gene for fertility restoration. The F2's of crosses cms Ramona 50 × D7465, cms Ramona 50 × D7466, and cms Ramona 50 × D7467 gave a ratio of 15 fertile to 1 sterile, indicating that two gene pairs in these three lines were responsible for the fertility restoration. The best of this group was D7467 which restored fertility fully after being crossed with cms Ramona 50 (T. timopheevi cytoplasm).

Relationship of Dwarfing and Hybrid Wheat Mechanisms1

The effects of dwarfing genes on the fertility of hybrid wheat (Triticum aestivum L. em Thell) were studied in 36 topcrosses. Three spring wheat cultivars with pollen fertility restorer genes for Triticum timopheevi cytoplasm were crossed with 12 near‐isogenic height lines of the spring wheat cultivars ‘INIA 66’ and ‘Norteno 67.’ The 36 F1,'s were used to pollinate a ‘Noroeste 66’ male sterile line.The source of major dwarfing segregation used in the near‐isogenic lines came from the cultivar ‘Olesen’ dwarf. Segregation for both height and restoration genes in a male‐sterile cytoplasm was studied. The degree of restoration and sterility was measured by the number of seeds set in one spike/plant. Frequency distributions revealed a larger‐than‐expected proportion of sterile plants in the short height groups and a larger‐than‐expected number of fertile plants in the tall height groups. Correlation studies in seed‐bearing plants showed a weak but positive relationship between plant height and fertility.

Basic Studies on Hybrid Wheat Breeding VI.Production of Male Sterile Lines of Common Wheat Cultivars

The male sterile T. timopheevi cytoplasm was introduced by successive backcrosses intcp 20 Japanese and 13 American cultivars of common wheat. In advanced backcross generations, all but four cultivars turned out to be complete or nearly complete male-steriles. Junrei-Komugi, and Norin 50, 52 and 69 were found to be carriers of a weak fertility restoring gene(s). From an investigation of their pedigrees, Junrei-Komugi, and Norin 50 and 52 were assumed to have received their gene(s) from their common ancestor, Shinchunaga, while Norin 69 appeared to have a different gene(s). Most male sterile lines of the other cultivars expressed male sterility constantly in different years (1969-1975), and in different localities (six locations from Kyushu ta Hokkaido in Japan). However, a few lines recovered selfed seed fertilities higher than 20% in Fukuyama in Chugoku District, where the environment seems to be shallow sterile for these lines. The out-crossing ability of all these male sterile lines was tested, using their seed setting rates under open-pollination. Male sterile lines of Aoba-Komugi, Bledsoe, and Lemhi 53 are considered to be good cross-fertilizers, while Karnvor, White Federation, Kokeshi-Komugi, Idaed 59, Fujimi- and Mikuni-Komugi are thought to be poor cross-fertilizers, the remainder being intermediate. With proper arrangement of pollinator rows, the male sterile lines tested showed a 51-83 seed-set by out-cross, which seems to be sufficient to guarantee hybrid seed production in Japan.

Pre-harvest sprouting in bread wheat (Triticum aestivum) as influenced by cytoplasmic male-sterility derived from T. Timopheevi

An investigation into the inheritance of pre-harvest germination (sprouting) in male sterile wheat is described. Sprouting does not occur in material which is hand emasculated prior to anthesis, but is present in cytoplasmic male-sterile lines. Thus sprouting appears to be associated with the interaction between the T. timopheevi cytoplasm and nuclei from T. aestivum. Different T. aestivum nuclei in T. timopheevi cytoplasm produce different sprouting tendencies.