Relationships between seed oil content and fatty acid composition in high stearic acid sunflower

The high stearic acid sunflower mutant CAS-3 is characterized by a low seed oil content, which might represent a constraint for the commercial production of high stearic acid sunflower oil. The objective of the present research was to investigate the relationships between fatty acid profile and seed oil content in CAS-3. Plants of CAS-3 were reciprocally crossed with plants of breeding line ADV-37, with high oil content and standard fatty acid profile. Oil content and fatty acid composition were measured in individual F₂ seeds and F₂ plants (F₃ seeds averaged). Both F₂ seeds and F₂ plants from the cross ADV-37 × CAS-3 had a significantly higher oil content than those from the reciprocal cross, which indicated the existence of cytoplasmic effects in the genetic control of the trait. A consistent negative correlation between oil content and palmitic acid and a positive correlation between oil content and oleic acid were detected both in F₂ seeds and F₂ plants. Conversely, no consistent correlation between oil content and stearic acid was observed, which suggested the feasibility of simultaneous selection for both traits.     

Genetic variability in plants regenerated from in vitro culture of sunflower (Helianthus annum L.)

In vitro regenerated sunflower (Helianthus annuus L.) plants (R₁) were self-fertilized and the R₂ generation was evaluated for qualitative traits. A broad range of phenotypic variation was observed and mutation frequencies were calculated. Some in vitro induced variant phenotypes were similar to known spontaneous or induced mutations in sunflower, while others were new. Chlorophyll and carotenoid deficiencies, chimaerical variegation, fasciated stem and capitulum, abnormal shoot development, and other morphological variations, were noted. Substitution of anthers with petaloid structures in a disk-floret mutant indicates a possible homeotic mutation induced by in vitro tissue culture.     

Novel variation for the tocopherol profile in a sunflower created by mutagenesis and recombination

α‐Tocopherol is the main tocopherol in sunflower seeds (>90%). Because it exerts a weak antioxidant action in vitro, its partial replacement by other tocopherols is an important breeding objective in this crop. The objective of this research was to develop novel tocopherol profiles in sunflower through mutagenesis and genetic recombination. Seeds of four ‘Peredovik’ accessions were used for chemical mutagenesis with ethyl methane sulfonate (EMS). Single‐seed screening in the M₂ generation resulted in two M₂ seeds, derived from different M₁ plants, with increased γ‐tocopherol contents of 19.2% and 96.7%, respectively. M₃ progeny from the M₂ seed with the 96.7% content bred true for high c‐tocopherol content, containing more than 90%γ‐tocopherol. M₃ progeny from the M₂ seed with only 19.2%γ‐tocopherol segregated in a range from 0 to 84.6%. Selection for high c‐tocopherol content produced an M_4:5 line, designated IAST‐1, with a stable high concentration of γ‐tocopherol. Crosses between IAST‐1 and T589, with an increased b‐tocopherol content, produced F₂ segregants with trans‐gressive levels of up to 77%β‐tocopherol or up to 68% d‐tocopherol. Both novel tocopherol profiles were confirmed in the F₃ generation.     

Inheritance of resistance to race 330 of Plasmopara halstedii in three sunflower lines

Sunflower lines RHA‐274, HA‐61 and RHA‐325 were studied for their resistance to race 330 of downy mildew (Plasmopara halstedii). The same inbred line, with normal (HA‐89) or sterile cytoplasm (cmsHA‐89) was used in all the crosses as susceptible parent, and, in each cross, only one genotype of the resistant parent was studied. The resistant‐to‐susceptible ratios obtained in the BC₁ and F₂ progenies from the crosses of the lines RHA‐274 and HA‐61 to cmsHA‐89 and HA‐89, respectively, suggested that, in each resistant line, two dominant genes are responsible for resistance to this downy mildew race. One of the genes (A) is epistatic to the other (B), and the recessive allele b in homozygosity is also epistatic to aa, with plants carrying aabb genotypes being resistant. Resistance to race 330 seemed to be controlled by two complementary genes in the sunflower inbred line RHA‐325, the dominant allele of one of them being present in cmsHA‐89. In the genotypes HA‐89 or cmsHA‐89, the existence of genes that modify the expected segregations following the crosses with resistant parents is proposed. It is concluded that, although major genes have been described as responsible for monogenic resistance to downy mildew, other types of regulation of this character, such as complementarity and epistatic relationships, do occur.     

Identification of genetic variation in Brassica napus seeds for tocopherol content and composition using near‐infrared spectroscopy technique

Tocopherol is an essential fat‐soluble nutrient for humans. Increasing the tocopherol content in Brassica napus seeds can add value to rapeseed vegetable oil; this has become an important breeding target. However, there is no efficient and non‐destructive method for selecting rapeseed accessions with high tocopherol contents. Here, we report the first near‐infrared reflectance spectroscopy (NIRS)‐based technique for predicting rapeseed tocopherol content. Individual seed tocopherol compositions were estimated from 373 rapeseed genotypes of different origins. This method and chemical methods produced comparable predicted values of the tocopherol constituents in the seeds. Three equations were generated for the prediction of tocopherol content by using a modified partial least squares (MPLS) model. The total tocopherol content for the determination coefficient of cross‐validation (R²_cv) (0.74), determination coefficient (RSQ) (0.76) and one minus the ratio of unexplained variance to total variance (1‐VR) (0.65) values indicates a strong correlation between the calibration and validation sets. Overall, our model confirmed the NIRS method as feasible for predicting tocopherol content in rapeseed and as an efficient screening tool for future breeding programs.     

Genetic control of in vitro-organogenesis in recombinant inbred lines of sunflower (Helianthus annuus L.)

Ninety-three recombinant inbred lines (F₈) of sunflower developed by the single-seed descent method from the cross ‘PAC-2 × RHA-266’ were used to screen their regenerability by organogenesis. The experiment was designed in randomized complete blocks with three replications. Each replication consisted of 10 Petri dishes with four explants. Cotyledons were excised from 2-day-old seedlings. Each cotyledon was divided into two pieces (four explants), which were incubated in solid regeneration medium consisting of full-strength Murashige and Skoog medium modified by adding hormones. A high genotypic variability for organogenesis parameters between genotypes was observed in this study. The difference between all recombinant F₈ lines and their parents was not significant, showing that the 93 inbred lines used in this experiment are representative of the total possible recombinant lines from the cross ‘PAC-2 × RHA-266′. These F₈ lines were not consciously selected for any trait; therefore, they represent a random set of lines segregating for the organogenesis parameters, as well as for other traits which could be important for breeding.     

Evaluation of different microsatellite motifs for analysing genetic relationships in cultivated sunflower (Helianthus annuus L.)

A total of 16 different simple sequence repeat motifs and an M13 repeat sequence were used as hybridization probes in order to examine the molecular relationships between two German inbred and eight North American sunflower lines of known pedigree, and to determine the suitability of the individual short tandem repeats for genetic analyses and their occurrence in the sunflower genome. Only the oligonucleotides (ACA)₆, (CAT)₆, (CATA)₅, (GACA)₄ and (GATA)₄ proved to be well suited for the generation of scorable fingerprinting patterns, while the other (microsatellite) sequences were either too abundant or too rare. Although different levels of polymorphism were present in hybridizations with the different simple sequence repeats (SSRs), a clear separation of the German material from the USDA Hnes was feasible not only by evaluating 20 probe-enzyme combinations, but also by combining every individual probe with four different restriction enzymes. In the American material, the expected relationships could be proved with the single exception of a line that did not group in accordance with its pedigree.     

Genetic analysis of tocopherol content and composition in winter rapeseed

The improvement of the nutritional value and the stability properties of rapeseed oil is partly hindered by the lack of information on the genetic control of tocopherols. The objectives of this investigation were to characterize the inheritance of tocopherols by using two factorial mating designs (North Carolina Design II, NC_II). The first (NC_II‐A) was produced with two sets of nine parents each, one selected for high and the other for low total tocopherol contents. In the second (NC_II‐B), each set consisted of six parents, which were either high or low for the α/γ‐tocopherol ratio. Parents and F₁ hybrids from both experiments were tested under field conditions in 1998 and 1999 using a completely randomized design with two replications. Only small differences were detected between mean values of parents and F₁ hybrids. General combining ability (GCA) variation in both experiments was highly significant (P < 0.01) for all tocopherol traits. Significant specific combining ability (SCA) effects were only detected for a‐tocopherol in NC_IIA and therefore for the α‐/γ‐tocopherol ratio. These results indicate that tocopherol contents and composition are controlled mainly by genes with additive effects. Interactions of F₁ hybrids and GCA effects with the environment were significant for only tocopherol contents and not for tocopherol composition.     

Genetic analysis of yield and related traits in sunflower (Helianthus annuus L.) in dryland and irrigated environments

The definition of a suitable breeding strategy in drought-prone environments is an important task for sunflower breeders. To achieve this task, reliable information on heritability and gene effects on yield and related traits under these conditions is necessary. Thirty six sunflower hybrids were produced by factorial cross of six male-sterile and six restorer lines. Parents and their hybrids were evaluated in eight environments. Six environments consisted of two adjacent trials in the experimental area, the first under irrigation and the second under dryland conditions, during 1987, 1988 and 1992. The other environments were: one early planting trial in dryland conditions, conducted during 1987, and a winter trial planted in January during 1988. Estimates of female variance (σ_f) were significant for seeds per head, seed weight, head sterile center, days to blooming and oil content. Female × male interactions (σ² _fm) were significant for all characters except harvest index and index of susceptibility to drought. Estimates of narrow sense heritabilities, calculated with information from analyses combined across environments, were 0.65 for yield, 0.80 for seeds per head, 0.84 for seed weight, 0.81 for head diameter, 0.60 for sterile head center, 0.72 for oil content, 0.61 for harvest index, 0.72 for biomass, 0.94 for days to bloom, and 0.42 for drought susceptibility index. Heritability estimates for individual environments showed more variation for yield than for other traits. Estimates for heritability of canopy temperature were high (0.68–0.79). Rainfed yield was positively correlated with yield components and negatively correlated with canopy temperature and susceptibility index. It is concluded that an efficient breeding strategy for sunflower under moderate drought-stressed conditions is the simultaneous selection for seed yield in both rainfed and irrigated environments together with selection for canopy temperature and stem diameter. This revised version was published online in July 2006 with corrections to the Cover Date.     

Genetic variation of tocopherol content in a germplasm collection of Brassica napus L.

The genetic variation for tocopherol contents was investigated in a very divergent collection of 87 winter rapeseed genotypes grown in the greenhouse and in two years in the field. Genotypic and environmental effects were highly significant for alpha-, gamma- and total tocopherol contents and the alpha-/gamma-tocopherol ratio. Field and greenhouse environments differed significantly (p < 0.001) for tocopherol traits, with greenhouse means up to 19% higher than field means. Alpha-tocopherol content ranged from 63 to157 mg kg^-1 seed, gamma-tocopherol content from 114 to 211 mg kg^-1 seed, total tocopherol content from 182 to367 mg kg^-1 seed, and the alpha-/gamma-tocopherol contents ratio from0.36 to 1.23. The resynthesized lines often have a remarkably low alpha-tocopherol content and consequently a low alpha-/gamma-tocopherol contents ratio. The lines with altered fatty acid composition displayed the highest variation for both tocopherol content and composition. No significant differences were observed among groups with different seed quality types [00], [0+] and [++]. Total tocopherol content was not correlated with the alpha-/gamma-tocopherol ratio, indicating that total tocopherol content is independent from tocopherol composition. Alpha- and gamma-tocopherol contents were also not correlated. Gamma-tocopherol contents showed a significant positive correlation with oil content (r = 0.34^**). This revised version was published online in July 2006 with corrections to the Cover Date.     

Inheritance and allelic relationships of fertility restoration genes for seven new sources of male-sterile cytoplasm in sunflower

The inheritance of fertility restoration of six mitomycin C and streptomycin‐induced cytoplasmic male‐sterile (cms) mutants and one cms line derived from Native American cultivar PI 432513 in sunflower was evaluated. These seven new cms sources were also compared with the commercially used cms PET1 (Helianthus petiolaris Nutt.) cytoplasm, using USDA inbred lines with restoration genes (Rf₁) specific for cms PET1 and new restoration lines identified for cms PI 432513. Restoration genes for cms PI 432513 were found in ‘Armavir’, VNIIMK, P21 and male‐fertile (MF) plants of PI 432513. F₂ and F₃ segregation ratios of crosses between cms PI 432513 and these restoration sources indicated a single dominant gene controlled fertility restoration. Progenies of cms PI 432513 testcrossed with F₁’s of half‐diallel crosses among the respective four homozygous restoration lines and RHA 274 suggested that the restoration genes of RHA 274, VNIIMK, P21 and PI 432513 were at the same locus. Restoration genes from VNIIMK, P21 and PI 432513 satisfactorily restored pollen stainability in the heterozygous condition. A very weak expression of the Rf gene in ‘Armavir’ was observed in the heterozygous condition. Fertility restoration capability of these genes for the six mutant cms HA 89 and cms HA 89 (in PET1 cytoplasm) was observed. The mutant cms HA 89 lines were also restored completely by RHA 266, RHA 274, RHA 280 and RHA 296, and F₂’s segregation ratios indicated single dominant gene control, implying a common cytoplasmic male sterility in all lines. F₁’s of half‐diallel crosses among RHA 266, RHA 273, RHA 274, RHA 280 and RHA 296 were testcrossed onto the cms lines, and their all MF progenies among lines, except RHA 280, confirmed that fertility restoration was controlled by a single Rf₁ gene locus. The restoration gene in confection line RHA 280, namely Rf₃, was at a different locus than Rf₁ and was equally capable of restoring all the cms lines. Cms HA 89 mutants and cms PI 432513 are in H. annuus cytoplasm, and are agronomically equal in hybrid performance to the cms PET1 used in commercial sunflower hybrids. These new cms lines will provide immediate alternative cms sources for reducing the genetic vulnerability resulting from the exclusive use of the single cms source PET1 in sunflower hybrid production.     

Inheritance of high palmitic acid content in the sunflower mutant CAS-12 and its relationship with high oleic content

CAS‐12 is a sunflower mutant with increased levels of palmitic (C16: 0 = 30%) and oleic (C18: 1 = 55%) acids in its seed oil, hence it has a reduced linoleic acid content (C18: 2 < 5%). This study was conducted to determine the inheritance of high C16: 0 content and its relationship with high C18: 1 content in CAS‐12. Reciprocal crosses involving CAS‐12, CAS‐5 (high C16: 0 content), HAOL‐9 (high C18: 1 content) and HA‐89 (standard fatty acid profile) were made. The F₁, F₂ and BC₁F₁ generations were obtained. The genetic control of the high C16: 0 trait in CAS‐12 was partially recessive and gametophytic. In all cases, this character segregated in the ratio 19: 38: 7 (low: intermediate: high C16: 0 content) in the F₂ generation. These results, together with the lack of segregation for C16: 0 content in crosses between CAS‐12 and CAS‐5, indicated that the genetic control of the high C16: 0 trait in CAS‐12 was similar to that in CAS‐5 in being controlled by partially recessive alleles (p1, p2, and p3) at three loci. Crosses between HA‐89 and CAS‐12, and HAOL‐9 and CAS‐5 (segregating for C16: 0 and C18: 1) demonstrated that the high C16: 0 and the high C18: 1 traits were independently inherited. However, C18: 1 segregation in these crosses exhibited reversal of dominance. Apparently, the low C18: 1 parental lines carried modifier genes causing the deviation.     

Dynamics of oil and tocopherol accumulation in sunflower grains and its impact on final oil quality

Tocopherols are one of the most important bioactive compounds in vegetable oils. It is known that these antioxidants present a dilution like relationship with oil weight per grain but the mechanism underlying this relationship are unknown. The aim of this work was to analyze the dynamics of tocopherol accumulation in sunflower grains, its relationship with oil accumulation and its effects on final oil quality in genotypes with different fatty acid composition. Three field experiments were conducted with genotypes with different potential fatty acid composition (a traditional, a high oleic and a high stearic−high oleic) and treatments with different source (intercepted solar radiation) or sink (grains) during grain filling to obtain varied grain filling conditions and grains with different oil concentration and oil unsaturation. Intercepted solar radiation modified oil per grain but did not affect tocopherol per grain. The rate of accumulation explained 79% and 74% of the oil and tocopherol per grain variation, respectively. When intercepted solar radiation increased, the duration of the period of oil and tocopherols accumulation increased, being the first the most responsive. These differences in the duration of accumulation periods are reflected in a larger relative increase in oil than tocopherols per grain and thus a dilution of the latter in the oil. These differences in the dynamics of oil and tocopherol accumulation are common to genotypes with different level of unsaturation. These results help to understand the mechanism associated with the dilution curve of oil tocopherol concentration reported in the literature.     

R-41, a sunflower restorer inbred line, carrying two genes for resistance against a highly virulent Spanish population of Orobanche cernua

The inheritance of resistance to a highly virulent population of sunflower broomrape (Orobanche cernua) has been studied in R-41, a Spanish sunflower restorer inbred line. Using the cytoplasmic male-sterile inbred line HA-89 (cms; very susceptible to this population of broomrape) as a female parent, progenies of the cross with R-41, i.e. Fl, F2 and BC1 to both parents, as well as the parental lines, were analysed for their reaction to the broomrape population EC-94. The goodness of fit of the observed vs expected segregation ratios indicated that the inheritance of resistance to broomrape in line R-41 is conferred by two independent dominant genes.     

Morphological and cytological study in a new type of cytoplasmic male-sterile line CMS-GIG2 in sunflower (Helianthus annuus)

Cytoplasmic male sterility (CMS) is essential for sunflower hybrid production. CMS-GIG2, a new sunflower CMS type, was further confirmed by crossing with the maintainer and restorer lines for the CMS-PET1, both of which maintain the male sterility of CMS-GIG2. Meiotic division in CMS-GIG2 was observed with 4'6-diamidino-2-phenylindole dihydrochloride staining, indicating that microspore formation was disrupted before the meiotic cytokinesis was completed. Light microscopy observation showed that both middle layer and tapetal cells expand radially rather than degrade over time, followed by failure to form tetrads and normal microspores. This morphological defect leading to male sterility in CMS-GIG2 differs from that observed in the PET1 CMS type. CMS-GIG2 will certainly provide additional genetic diversity for sunflower hybrid breeding programmes.     

Pure line selection for improved yield, oil content and different fatty acid composition of sesame, Sesamum indicum

The aims of this study were estimation of the variability of seed yield, oil content, oleic acid and linoleic acid content in single plant progenies and lines derived from Turkish sesame populations, and the improvement of different type lines with regard to high seed yield, high oil content and specific fatty acid composition. Different types of lines were developed—i.e. lines which yielded over 1000 kg seeds/ha from the yield type progenies and lines which yielded over 63% of oil content from oil type progenies—but it was not possible to improve lines where fatty acid composition differed dramatically from that of oleic and linoleic type progenies.     

Thresholds for leaf expansion and transpiration response to soil water deficit in a range of sunflower genotypes

The sunflower (Helianthus annuus L.) crop in southern Europe suffers from intense and frequent periods of water deficit. Minimisation of water loss in response to water deficit is a major aspect of drought tolerance and can be achieved through the lowering of either leaf area expansion rate or transpiration per unit leaf area (stomatal conductance). During three greenhouse pot experiments, leaf expansion (LE) and transpiration (TR) rates were monitored as the soil dried progressively for about 15 days. This study aimed to quantify the response of these two physiological processes to water deficit, expressed as the fraction of transpirable soil water (FTSW): response thresholds (onset of decline) were estimated on 25 sunflower genotypes from different generations of selection history. From these relationships, the thresholds below which LE and TR started to decrease from the control were calculated: little change was observed until FTSW was close to 0.6 for leaf expansion and 0.4 for daily transpiration. Variability in the response of all genotypes for expansion and transpiration control was better described using specific thresholds for each genotype rather than generic thresholds for sunflower. The ranking of genotypes was found to be unaffected for transpiration rate control, but this was not the case for leaf expansion. Identified response thresholds were not mutually correlated, suggesting that sunflower controls leaf expansion and transpiration rate independently. Neither was correlated with the release date of the genotype, suggesting that these traits were not subject to selection within evaluation environments.     

Breeding for Sclerotinia resistance in sunflower: A modified screening test and assessment of genetic variation in current germplasm

Sclerotinia sclerotiorum (Lib.) de Bary is one of the most devastating pathogens of sunflower (Helianthus annuus), causing yield losses up to 100%. This study was conducted to (1) screen sunflower lines for their reaction to Sclerotinia using artificial infection by a modified leaf test, and (2) identify the most suitable trait for screening against Sclerotinia resistance. A set of 85 inbred lines was screened in three environments by infecting the leaves with mycelium and covering them with plastic bags containing water. Susceptibility was assessed by leaf lesion, petiole score, stem rating, stem lesion and diseased stems. The test proved to be effective and practicable, with an infection rate of up to 99%. All scoring traits were strongly correlated with each other and effective in differentiating among susceptible and resistant lines. Although petiole score had the highest heritability, its association with leaf length hampers its utilization as a scoring trait. Stem lesion was a good predictor of mid-stalk rot and can be recommended as a reliable trait for screening sunflower lines against Sclerotinia based on its high heritability estimate (h²= 0.59) and close correlations with other traits measured after artificial and natural infection.