The mutagenic effect of treatment with EMS at different temperatures in Pisum sativum

Dry peas were treated with solutions of 0.13% and 0.2% EMS for 24 hours at temperatures of 6°, 12°, 18° and 24°C, respectively. The effect of the temperature during treatment was studied in the M₁ and in the M₂ generation.The germination capacity of the treated seeds and the percentage surviving M₁ plants were unfavourably affected only after treatment at 24°C. Leaf spotting and growth inhibition in the M₁ plants increased with increasing temperature, while fertility decreased. After treatment at 24°C more than 75% of the harvested M₁ plants were completely sterile.With rising temperature the percentage mutants in the M₂ increased from 8 to 50%. However, the fertility of the mutants was considerably reduced. The sterility after treatment with EMS, therefore, must be at least partly due to genetic changes. This sterility greatly diminishes the mutagenic efficiency of EMS treatment at high temperatures.     

EMS Induction of Early Flowering Mutants in Spring Rape (Brassica napus)

Studies were undertaken to induce early flowering mutants by ethyl methanesulphonate (EMS) treatments of Brassica napus seeds. EMS treatments for 12 h of a highly inbred B. napus line TBS had adverse effects on M¹ plant development and fertility only when concentrations were greater than 1%. However, an EMS concentration of 1.5% did not reduce M₁ plant fertility to an extent which significantly reduced production of M₂ seeds. Genetic changes induced by EMS treatment and affecting flowering time were of three main types: (1) Changes within a polygenic system reflected by increased variation in flowering time among M₂ families. As the increase in variation was due primarily to a higher frequency of later flowering plants, these polygenic changes would be of little value in developing better-adapted cultivars. (2) Induction of a recessive mutation at a major gene locus which caused M₃ plants homozygous for the mutant gene to flower at least 20 days earlier than the parental line TB8. (3) Induction of a dominant mutation at a major gene locus which affects flowering time by causing a substantial reduction in vernalization requirement. M₂ plants carrying the mutant gene flowered as early as 59 days before the parental line. These major gene mutations could be rapidly exploited in the development of agronomically superior cultivars for short-season, lower rainfall environments in Western Australia.     

Mutation research in peas

Dry seeds of the pea variety Pauli were irradiated with 15,000 r X-rays, 18,000 r X-rays, 18,000 r rays and with neutrons; the latter treatment for 6 hours. In addition the chemical mutagen ethyl methane sulphonate (EMS) was used, in concentrations of 0.33% and 0.50% respectively.After raising the germlings in a glasshouse surviving plants were planted in the field. The germination capacity was only little disturbed after the irradiation; however, it was greatly reduced after the EMS treatment. A very large number of germlings did not grow further, their roots showing a deep brown discoloration, ultimately leading to death. Several other influences after planting in the field killed many other plants.The fertility of the M₁-generation was only little disturbed by the irradiation, but much by the EMS-treatment. In the M₂-generation, both after irradiation and after EMS-treatment, at higher dosages and concentrations lower percentages of mutated progenies were found than at lower dosages and concentrations. This was presumably due to the fact that the number of plants per M₂-line in the former case was on average twice as high. The percentage progenies with mutations and the percentage mutated plants was larger after EMS-treatment than after irradiation.Of normal looking plants with a good fertility, progenies were raised in non-replicated small plots. The average yield per plant and the distribution in the irradiations did not differ significantly from that of the untreated material. After EMS-treatment the average yield was significantly lower and the distribution significantly higher. The fertility disturbance caused by EMS, therefore, also became manifest in the M₃-generation.     

Moricandia arvensis cytoplasm based system of cytoplasmic male sterility in Brassica juncea: Reappraisal of fertility restoration and agronomic potential

Cytoplasmic male sterility (CMS) system based on the cytoplasm from Moricandia arvensis (mori) was investigated for fertility restoration and agronomic potential. Fertility restorer gene for mori CMS was introgressed from cytoplasm donor species as all the evaluated Brassica juncea genotypes (155) acted as sterility maintainers. The allosyndetic pairing between M^a and the A/B genome chromosomes in the monosomic addition plants (2n= 18II + 1M^a) facilitated the gene introgression. Partial fertility restoration (43–52% pollen grain stainability) in F₁ hybrids and absence of segregation for male sterility in F₂ progenies suggested gametophytic control of fertility restoration. The pollen fertility in the F₁ hybrids was, however, sufficient to ensure complete seed set upon bag selfing. Introgression from M. arvensis also helped in correction of chlorosis associated with mori cytoplasm in CMS and fertile alloplasmic B. juncea plants. Yield evaluation of thirty F₁ hybrids having the same nuclear genotype but varied male sterilizing cytoplasms (mori, oxy, lyr, refined ogu), in comparison to respective euplasmic hand bred control hybrids, allowed an estimate of yield penalty associated with different CMS systems. It ranged from 1.8% to 61.6%. Hybrids based on cytoplasmically refined ogu were most productive followed by those based on cytoplasmically refined mori CMS. The male sterility systems emanating from somatic hybridization were found superior than those developed from sexual hybridization.     

Improvement of seed yield in Vicia faba L. By using experimental mutagenesis. II. Comparison of gamma-radiation and ethyl-methane-sulphonate (EMS) in production of morphological mutants

Summary Gamma-rays and EMS have been used in two dosages to test their effectiveness in inducing mutations in one major (cv. Aguadulce) and one minor (cv. Manfredini) Vicia faba cultivar. The plants of the M₁ and M₂ generations showed reduced emergence, survival, and fertility. EMS was more effective than gammarays in inducing mutations giving a mutation frequency in M₁ and a frequency of M₂ mutants from two to four times higher. The two mutagenic agents do not seem to differ in mutation spectrum. The minor cv. Manfredini was more sensitive to the lower dosage of gamma-rays than the major cv. Aguadulce. Both mutagens proved to be effective in enlarging the morphological variation of both cultivars.Research work supported by C.N.R. special grant I.P.R.A.-Sub-project 1. Paper N. 515.     

A partially male-sterile mutant line of soybeans,Glycine max (L.) Merr.: Characterization of the msp phenotype variation

Summary Investigations of variable expression of msp partial male sterility in soybeans (Glycine max (L.) Merr.) showed that higher temperatures promote male fertility in msp homozygotes and showed that infectious agents are not intrinsic to the sterility system. Exchange grafts failed to modify fertility levels of msp msp rootstocks, Msp Msp scions, and their self-progeny. Tests for soybean mosaic virus and tobacco ringspot virus were negative in partially male-sterile plants, in control fertile plants, and in self-progeny of grafted plants. Growth-chamber experiments and field observations manifested that male fertility of msp msp plants is higher in hot environments than in cooler ones. The unexpected aberrant ratios of fertile to partially male-sterile plants observed in 1977 (Stelly & Palmer, 1980) are explainable on the basis of msp temperature sensitivity.Our observations suggest that homogeneous msp msp populations may be increased in hot environments.Research Geneticist, SEA-AR, USDA, Iowa State University, Ames, Iowa 50011, USA.     

The increase of seed fertility of Brassicoraphanus through cytological irregularity

Summary Amphidiploids (Brassicoraphanus) were produced by means of colchicine treatment of F₁ hybrids between Brassica japonica Sieb. and Raphanus sativus L. The cytology of the amphidiploids was studied from F₁ to F₃ generations. Some plants had the euploid chromosome number 2n=38, whereas others had the aneuploid number 2n=37. One or two of either quadrivalents or trivalents, as well as some univalents, were seen in most of the plants examined. All the plants showed a low seed fertility. In F₃ generation there arose some yellow-flowered plants, all of which showed a higher seed fertility than normal white-flowered plants. It is postulated that the change of flower colour might originate in the segmental exchange of only partially homologous chromosomes following multivalent formation. A gene causing white flower colour was perhaps closely linked to a gene causing sterility, and both genes were probably excluded together through the segmental exchange of the chromosomes. Therefore, it can be said that the increase of fertility was induced by cytological irregularity.     

EMS-induced genic male sterility in Arabidopsis thaliana: A model selection experiment

Summary Seeds of Arabidopsis thaliana were treated with EMS to initiate a small-scale selection program for genic male sterility. Six lines of independent origin were obtained. Of these, four were discontinued in M₅, as they segregated incompletes, and two are maintained by full-sib mating, viz. line 97, which shows complete expression throughout the inflorescence, and line 112, which shows complete expression except in the later flowers. At the same time the additional aims were reached of unreduced female fertility and perfect phenotypic resemblance to the original line. The six mutations arose at five different loci. An attempt to extract a cytoplasmic male sterile failed.

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The mutual independence of M1-fertility and mutant yield in EMS treated tomatoes

After EMS treatment of tomato seeds, a positive correlation was found between degree of M₁-sterility and percentage of seedling mutants in M₂. However, when the treated seeds were grouped according to degree of germination delay, a within-day correlation was present only in the group of earlier germinators. An explanatory model is proposed which implies that the sporocyte tissue of a cluster (or part of it) may either derive from a more sensitive (G₁) initial in the dormant embryo, or from a less sensitive one (G₂), and that within a population of clusters derived from cells at the same level of overall sensitivity, the correlation is absent. For practical mutation breeding the implication is, that breeding efficiency can be increased by harvesting only sectors with good fertility since this does not lead to loss in mutant yield. A mutagenic treatment should be such that fully fertile M₁-sectors are not too rare.This research was part of a project resulting from a contract between The National Council for Agricultural Research T.N.O., the Agricultural University, and the Association EURATOM-I.T.A.L. at Wageningen.Part of the data were presented as a short communication (43) at the Symposium on the Mutational Process (Praha, August 9–11, 1965).     

Development of cytoplasmic-genic male sterility in safflower

An interspecific cross was made between Carthamaus oxyacantha and the cultivated species C. tinctorius to develop a cytoplasmic‐genic male sterility (CMS) system in safflower. C. oxyacantha was the donor of sterile cytoplasm. The 3: 1 segregation pattern observed in BC₁F₂ suggested single gene control with dominance of male‐fertility over male‐sterility. The information obtained from crossing male sterile X male fertile plants in BC₁F₃ and BC₁F₄ generations showed statistically significant single gene (1: 1) segregation for male sterility vs. male fertility. The results demonstrated that C. tinctorius possesses a nuclear fertility restorer gene and that a single dominant allele restored fertility (Rf) in progeny carrying CMS cytoplasm of C. oxyacantha. Male sterility occurred with the homozygous recessive condition (rfrf) in a sterile C. oxyacantha cytoplasm background and not in the normal cytoplasm of C. tinctorius. The genetic background of different restorer lines of C. tinctorius having normal cytoplasm did not effect fertility restoration. The absence of male sterile plants in C. tinctorius populations ruled out the possibility of genetic male sterility. Normal meiosis in F₁ and BC₁F₂ ruled out a cytogenetic basis for the occurrence of male sterility.     

An interspecific cross in Cucurbita: C. lundelliana bailey ×C. maxima duchesne

F₁, F₂, and backcross progenies derived from an interspecific cross between Cucurbita lundelliana and C. maxima were analyzed by using the hybrid index method of Anderson. The results suggest that C. lundelliana is dominant for most characters. There is a good deal of sterility, particularly in the backcross to C. maxima. It appears feasible, however, to transfer such characters as powdery mildew resistance, perennial habit, solid placenta, etc. from C. lundelliana to C. maxima by conventional plant-breeding techniques. The mean pollen fertility for the F₁, F₂ and backcross to C. lundelliana was respectively 17.6%, 28.4% and 5.2%: for the backcross to C. maxima. 65.9%.     

Cytomorphological studies in an intergeneric hybrid between Gossypium hirsutum L. (2n=52) and Hibiscus panduraeformis Burm.

Summary An intergeneric hybrid (2n=38) between Gossypium hirsutum L. (2n=52) × Hibiscus panduraeformis Burm. (2n=24) was obtained by pollinating about 2000 flower buds of G. hirsutum var. Gregg Male Sterile with pollen from H. panduraeformis. The F₁ hybrid was intermediate in plant habit, but possessed gossypol glands and nectaries on the leaves, bolls containing seeds with fuzz and lint as dominant characters of G. hirsutum. Flowers with yellow corolla and anthers; purple petal spot, profuse growth of epidermal hairs on all plant parts including the boll sutures, and jassid tolerance were dominant characters of H. panduraeformis. The partial fertility of the F₁ indicated the possibilities of combining jassid and drought tolerance of H. panduraeformis with the desired economic characters of G. hirsutum for rainfed cultivation.The F₁ hybrid showed various meiotic irreguarities and about 40% pollen sterility. Formation of the normal bivalents occurred quite frequently suggesting a close relationship between the parental species. The sterility observed in the hybrid may be due to small structural differences between the chromosomes of the two genera and meiotic abnormalities.     

Dwarf-twisted: An induced mutation in Triticum durum Desf

Summary A mutant phenotype, mainly characterized by undulated leaf surface, dwarfism and sterility, has been repeatedly induced by physical (X-rays; thermal and fast neutrons) and chemical (diethylsulphate and ethylmethanesulphonate) mutagenic agents in different varieties of durum wheat. This phenotype is easily detectable at the seedling stage. Neutron treatments are the most effective method to induce this phenotype and a maximum of 0.6% mutations per M₁ spike-progeny was induced after the highest dose of thermal neutrons. On the average 0.23 mutations per 100 M₁ spike progenies and 0.55 mutant seedlings per 1000 M₂ plants were found after all treatments.Genetic analysis of different dwarf-twisted phenotypes shows that this character is conditioned by one recessive gene with an average segregation ratio of 22.9%. As the dwarf-twisted phenotypes segregate in highly fertile M₁ spikes and no evident chromosome changes have been detected in this caryotype, the hypothesis of a point mutation has been proposed.Contribution no. 171 from the Laboratorio Applicazioni in Agricoltura del C.N.E.N., C.S.N. Casaccia, S. Maria di Galeria, Roma, Italy.     

A unique introgression from Moricandia arvensis confers male fertility upon two different cytoplasmic male-sterile lines of Brassica juncea

A Brassica juncea line carrying an introgression from Moricandia arvensis restored male fertility to two cytoplasmic male‐sterile (CMS) B. juncea lines carrying either M. arvensis or Diplotaxis catholica cytoplasm. Genetics of fertility restoration was studied in the F₁, F₂, F₃ and backcross generations of the cross between CMS and fertility‐restorer lines. No male‐sterile plants were found in F1‐F₃ generations of the cross between CMS [M. arvensis] B. juncea and the restorer. However, a 1: 1 segregation for male sterility and fertility was observed when the F1 was pollinated with non‐restorer pollen from a euplasmic line. These results clearly show that restoration is mono‐genic and gametophytic. In CMS lines carrying D. catholica cytoplasm, the restorer conferred male fertility to the F1 and showed 3: 1 and 1: 1 segregations for male fertility and sterility in F₂ and BC1 generations, respectively, indicating a monogenic, sporophytic mode of fertility restoration. The results were also supported by pollen stainability in the F1 which was about 65% in M. arvensis‐based CMS and >90% in D. catholica‐based CMS. The above results are discussed in the light of previous molecular studies which showed association between CMS and atpA in both systems.     

An induced brachytic mutant of chickpea and its possible use in ideotype breeding

Mutations were induced in chickpea (Cicer arietinum L.) cultivar ‘JG 315’ through treatment of seeds with ethyl methane sulphonate (EMS). One of the mutants, named JGM 1, had brachytic growth (compact growth), characterized by erect growth habit, thick and sturdy stem, short internodal and interleaflet distances and few tertiary and later order branches. It was isolated from M₂ derived from seeds treated with 0.6% EMS for 6 h. Segregation analyses in F₂ progenies of its crosses with normal chickpea genotypes (JG 315, ICC 4929, and ICC 10301) suggested that a single recessive gene controlled brachytic growth in JGM 1. This gene was not allelic to the br gene for brachytic growth in spontaneous brachytic mutant E100YM. Thus, the gene for brachytic growth in JGM 1 was designated br2 and the br gene of E100YM was redesignated br1. Efforts are being made to use JGM 1 in development of a plant type with short internodes and erect growth habit. Such plant type may resist excessive vegetative growth in high input (irrigation and fertility) conditions and accommodate more plants per unit area.     

Genetic and cytological analysis of a new spontaneous male sterility in radish (<Emphasis Type="Italic">Raphanus sativus</Emphasis> L.)

A male sterile plant appeared in the radish breeding program at the Hubei Academy of Agricultural Sciences, Hubei, China. In its progeny, a two-type (half of plants male sterile, the other half male fertile) line 01GAB was established. An F₂ population of 260 plants from a cross of male-sterile 01GAB and a male fertile line 9802H segregated for male fertility in a 3:1 ratio indicating that fertility was restored by a single dominant gene, here designated RsMs. A PCR-based DNA marker specific to the male fertility Rfob gene in 9802H was absent in 01GAB. Linkage analysis placed the RsMs locus 10.7 cM away from the Rfo locus. In an F₂ population of hybrids between 01GAB and male fertile 9802B, a co-dominant DNA marker for the RSultr3.2A (a radish sulfate transporter gene) locus was linked to the RsMs locus at 1.5 cM suggesting that fertility restoration in 01GAB was located in the region with known male sterility restorers in radish. However, no maintainer for the 01GAB source of male sterility has been identified so far. Cytological observations have shown that the abnormalities in male sterile anthers first appeared in tapetum at the tetrad stage, followed by a hypertrophy of the tapetal cells at the vacuolate microspore period. These results suggest that male sterility in 01GAB is likely to be genetic in nature, or it may represent a new type of the cytoplasmic male sterility.     

Tournefortii male sterility system in Brassica napus. Identification, expression and genetic characterization of male fertility restorers

A germplasm collection of 152 diverse rapeseed accessions from Canada, China, France, India, Poland and South Korea was assayed for identifying new fertility restorers and sterility maintainers for a Tournefortii (tour) cytoplasmic male sterility (CMS) system in rape‐seed. Only 16 (10.5%) genotypes showed complete fertility restoration following hybridization with tour CMS line NE 409A. Notable among these were GSL 8851, GSL 8953, Mokpo # 9, Mali, Buk‐wuk‐13, Kuju‐27 and Mokpo # 84. As many as 78 (51.3%) genotypes were perfect maintainers of sterility, the remaining 58 (38.2%) genotypes were classified as partial maintainers. To study the inheritance of fertility restoration, 20 CMS (tour) rapeseed lines were crossed with the four best fertility restorers, namely GSL 8851, GSL 8953, Kuju‐27 and Mokpo # 9, to obtain F₂ and test cross populations. Segregation data indicated that fertility restoration for tour CMS was governed by two genes, of which, one is stronger than the other (χ²_12:3:1). Differences in gene interactions were also observed (χ²_9:3:4) which could be explained on the basis of influence of female parent genotypes/or modified expression of the restorer gene(s) in different genetic backgrounds. Tests of allelism indicated that the restorer genes present in the four restorers evaluated were allelic.     

Interspecific hybridization in the genus Nicotiana involving a new Australian species N. umbratica Burbidge

Summary The F₁ inter-subgeneric hybrid N. glutinosa L. × N. umbratica Burbidge revealed that N. glutinosa (2n=24) is distantly related to N. umbratica (2n=46). The synthetic amphiploid N. glutinosa × umbratica (2n=70) which is stable in morphology and cytology, with good fertility for five generations, is an experimental verification of the postulate by Goodspeed (1954).     

Cytoplasmic and cytogenetic relationships among tetraploid Triticum species

Summary The F₁ hybrids from crosses of 59 accessions of wild and cultivated Triticum types including amphidiploids T. boeoticum-Ae. squarrosa, T. timopheevi-Ae. squarrosa, T. timopheevi-T. monococcum, T. boeoticum (4n), T. macha, and T. Zhukovskyi with T. durum Sel. 56-1 and/or T. aestivum were examined for male sterility and chromosome pairing at metaphase I of meiosis in pollen mother cells. Those hybrids which produced male-sterile F₁'s were recurrently backrossed with pollen from T. durum or T. aestivum to study segregation for male sterility and/or to confirm cytoplasmic male sterility.All T. timopheevi and T. araraticum accessions and several T. dicoccoides types, including T. dicoccoides var. nudiglumis from the Turkey-Iran-Iraq area, had male sterility inducing cytoplasm. The chromosome pairing in the F₁ hybrids indicated that all tetraploid Triticum accessions with male sterility inducing cytoplasm had genome AAGG. T. dicoccoides Körn types from the Turkey-Iran-Iraq area had genomes AABB and did not have male sterility inducing cytoplasm. Therefore, T. dicoccoides Körn and the T. timopheevi complex differ from each other cytoplasmically and cytogenetically and occur sympatrically in the Turkey-Iran-Iraq area.Possibly, the cytoplasm of the emmers was not derived from the putative diploid progenitors, T. boeoticum, Ae. speltoides, or Ae. bicornis as indicated by their nucleo-cytoplasmic and cytogenetic relationships with the tetraploid Triticum species. The cytoplasmic differences among Ae. speltoides, T. araraticum and T. timopheevi are of a relatively smaller magnitude than the cytoplasmic differences among T. timopheevi, T. boeoticum, and the emmers. A complete analysis of nucleo-cytoplasmic relationships among Triticum and Aegilops species may indicate the cytoplasmic donor(s) to the two tetraploid Triticum species complexes.Authorized for publication 19 July, 1972 as Paper No 397 in the Journal Series of the North Dakota Agricultural Experiment Stations.     

Cytoplasmic homology between Aegilops mutica Boiss. and Ae. ovata L.

Summary Triticum aestivum L. em Thell. (2n=42; AABBDD), and T. durum Desf. (2n=28; AABB) genomes were substituted into the cytoplasms of Aegilops mutica Boiss. (2n=14; M_tM_t), Ae. heldreichii Holzm. (2n=14; MM), Ae. uniaristata Vis. (2n=14; M^uM^u), and Ae. ovata L. (2n=28; C^uC^uM^oM^o), to identify the M-genome diploid cytoplasm donor of Ae. ovata. Substitution of the T. durum genome into Ae. uniaristata cytoplasm resulted in a large proportion of shriveled inviable seeds. A few plump viable seeds were obtained all of which produced male-sterile plants having one univalent or telocentric chromosome from Ae. uniaristata. The T. aestivum plants having Ae. uniaristata or Ae. mutica cytoplasms were fertile. However, Ae. mutica was similar to Ae. ovata in the induction of delayed maturity and tall robust growth habit to the T. durum and T. aestivum plants. Cytoplasms of the other C- and M-genome diploids Ae. umbellulata Zhuk. (2n=14; C^uC^u) and, Ae. heldreichii (2n=14; MM) earlier had been shown to differ from that of Ae. ovata. Therefore, Ae. mutica is the most likely cytoplasm and M-genome donor to Ae. ovata.