Sunflower Inbred Lines Research Articles

Validation of a method for the analysis of phytosterols in sunflower seeds

AbstractPhytosterols are natural compounds that contribute to lower serum cholesterol in humans. Sunflower seeds and oils are rich sources of phytosterols. Breeding for phytosterol content in sunflower has been scarce thus far, mainly because of the lack of analytical methods suitable for use in plant breeding. The objective of this research was to validate a method for the analysis of phytosterols in small seed samples of sunflower. Samples consisting of six seeds were analyzed for phytosterol content in a set of 87 inbred lines using a method adapted to small samples. The accuracy of the method was evaluated through the standard error of the analysis of replicates of ground samples, which was 72.12 mg/kg compared to average values of 1665.3 and 1887.2 mg/kg seed in the samples. Sunflower inbred lines showed ranges of variation from 1426.0 to 4710.0 mg/kg seed and from 2855.2 to 9752.0 mg/kg oil. The method correlated strongly with the conventional method based on the analysis of extracted oils (r = 0.85). The results indicated that analysis of phytosterols on samples consisting of sunflower seeds is an accurate approach for breeding and genetic studies, in which extraction of the seed oil is not feasible.Practical applications: Phytosterols are usually analyzed in extracted oils. However, studies in plant breeding and plant sciences often require a direct analysis of phytosterols in seeds, without previous oil extraction (e.g. large‐scale screening of germplasm in breeding programs or genetic studies). Our results will be useful for plant scientists interested in the analysis of phytosterols in small samples of plant tissues.

Relationship between virulence and aggressiveness in Plasmopara halstedii (sunflower downy mildew)

Relationship between virulence and aggressiveness was studied in seven Plasmopara halstedii (sunflower downy mildew) pathotypes including five progeny pathotypes of races 300, 304, 314, 704 and 714 arising from two parental pathotypes of races 100 and 710. Aggressiveness criteria including percentage infection, latent period, sporulation density and reduction of hypocotyl length were analysed in one sunflower inbred line showing a high level of quantitative resistance. There were significant differences between P. halstedii pathotypes for all aggressiveness criteria. Pathogenicity of progeny pathotypes as compared with parental ones (relationship between virulence and aggressiveness) seems to be positive, negative or uncorrelated. Hypothesis explaining these cases are discussed.

Can percentage infection and dwarfing be used to differentiate aggressiveness in Plasmopara halstedii?

Aggressiveness was studied in seven Plasmopara halstedii (sunflower downy mildew) parental isolates of races 100, 300, 304, 314, 704, 710 and 714 using five single zoosporangium isolates per parental isolate. Aggressiveness criteria, including percentage infection and dwarfing (reduction of hypocotyl length), were analysed in one sunflower inbred line showing a high level of quantitative resistance. Analysis of five single zoosporangium isolates of each parental isolate showed variability within parental isolate for the two aggressiveness criteria, but not for all parental isolates for percentage infection and vice versa for the reduction of hypocotyl length. Percentage infection showed high values irrespective of the parental isolate used. However, all the parental isolates caused a large reduction in seedling size except for the isolate of race 314. Although percentage infection and reduction of hypocotyl length could be used to differentiate aggressiveness in P. halstedii, it seems that these criteria played a limited role to define P. halstedii isolates according to their aggressiveness.

Relationship between aggressiveness and zoosporangia viability in Plasmopara halstedii (sunflower downy mildew)

Relationship between aggressiveness and zoosporangia viability was studied in seven Plasmopara halstedii (sunflower downy mildew) isolates of races 100, 300, 304, 314, 710, 704 and 714. Aggressiveness criteria including latent period and sporulation density were analysed on sunflower inbred line showing a high level of quantitative resistance. There were significant differences between pathogen isolates for the two aggressiveness criteria. Viability analyses were performed on oval and spheric zoosporangia. The number of zoospores released from oval zoosporangia was significantly higher than those released from spheric ones. The oval zoosporangia for more aggressive isolates of races 100 and 3xx produced more zoospores than the oval ones for less aggressive isolates of races 7xx. There was a significant correlation between aggressiveness criteria and the number of zoospores released from oval zoosporangia and vice versa for zoospores released from spheric ones. It is concluded that the relationship between aggressiveness and oval zoosporangia viability may be established in P. halstedii.

Pathogenic variability in Plasmopara halstedii (sunflower downy mildew)

Pathogenic variation was studied in 19 Plasmopara halstedii isolates belonging to races 100, 300, 710 and 714. Based on the reaction for the P. halstedii isolates to four sunflower hybrids H1 to H4 differing only in their downy mildew resistance genes, there were differences in virulence spectrum in pathogen isolates. All sunflower hybrids were resistant to isolates of races 100 and 300 and sensible to isolate of the race 714, H1 was only sensible to isolates of race 710. Regarding aggressiveness criteria on two sunflower inbred lines showing different levels of quantitative resistance, there were significant differences in aggressiveness for P. halstedii isolates. The index of aggressiveness (sporulation density/latent period) varied between 8.1 and 23.6. There was a significant correlation (r = −0.456) between the two aggressiveness criteria at p < 0.05.

Aggressiveness in Plasmopara halstedii (sunflower downy mildew)

Aggressiveness was studied in seven Plasmopara halstedii (sunflower downy mildew) pathotypes: 100, 300, 304, 314, 704, 710 and 714. Aggressiveness criteria including percentage infection, latent period, sporulation density and reduction of hypocotyl length (dwarfing) were analysed in one sunflower inbred line showing a high level of quantitative resistance. Genetic relationships were detected between the seven pathotypes using 12 EST-derived markers. Pathotypes 100, 300, 304 and 314 were characterized with shorter latent period and higher sporulation density than pathotypes 710, 704 and 714. All pathotypes showed high percentage infection values and caused a large reduction in seedling size except for pathotype 314 involved in dwarfing. Pathotypes 714, 704 and 314 had an intermediary genetic position between the pathotypes 100 and 710. No correlation was detected between aggressiveness traits and EST genotypes.

Selection of a Sunflower Line with Multiple Herbicide Tolerance That Is Reversed by the P450 Inhibitor Malathion

Ninety-seven inbred lines of sunflower were screened in the field by treatment with a combination of imazamox and malathion, an inhibitor of cytochrome P450 monooxygenases (P450s), to identify sunflower lines with natural tolerance to the herbicide reversed by malathion. One tolerant line, named TolP450-1, was selected and characterized in the field and in the greenhouse to evaluate its response to the herbicides imazamox, prosulfuron, and atrazine at different plant development stages (germination, emergence, and third difoliate) with and without malathion. For all herbicides and all development stages analyzed, TolP450-1 showed significantly higher tolerance compared with the susceptible line RHA266. In all cases, the tolerance was reversed by malathion. This sunflower line, tolerant to multiple herbicides, may be useful in helping to manage herbicide-resistant weeds by allowing additional herbicides to be used in this oilseed crop.

Genetic dissection of tocopherol and phytosterol in recombinant inbred lines of sunflower through quantitative trait locus analysis and the candidate gene approach

Sunflower (Helianthus annuus L.) contains tocopherol, a non-enzymatic antioxidant known as lipid-soluble vitamin E, and phytosterol, with interesting properties, which can result in decreased risk of chronic diseases in humans and with several beneficial effects in plants. The genetic control of tocopherol and phytosterol content in a population of 123 recombinant inbred lines of sunflower was studied through quantitative trait loci (QTL) analysis using 190 simple sequence repeats and a gene-based linkage map. Seven experiments were conducted in different environments in France and Iran during 2007 and 2008. Each experiment consisted of three replications. Means over all environments were used for QTL mapping. Five QTL for total tocopherol content on linkage groups 1, 8, 10 and 14 accounted for 45% of phenotypic variation, whereas four QTL for total phytosterol content on linkage groups 1, 2, 16 and 17 explained 27% of the phenotypic variation. GST, PAT2, SFH3 and POD genes showed co-localization with QTL for total phytosterol content. SMT2 is also mapped on linkage group 17 near the QTL of total phytosterol content. Four candidate genes, VTE4, HPPD, GST and Droug1, exhibited co-localization with QTL for total tocopherol content. The candidate genes associated with tocopherol and phytosterol, especially HPPD, VTE4 and SMT2, could be used for alternation of the tocopherol and phytosterol content of sunflower seeds through the development of functional markers.

General and Specific Combining Ability for Quantitative Characters in Sunflower

Seven sunflower (Helianthus annuus L.) inbred lines were crossed in 7´7 half diallel to obtain 21 F1 hybrids which further used to estimate general combining ability (GCA) and specific combining ability (SCA) effects for yield, head diameter, 1,000-seed weight, plant height and oil content of sunflower. The 7 inbred lines and 21 hybrids were planted in a randomized complete block design with three replications at Nakhon Ratchasima, Thailand during 2008-2009. General combining ability and specific combining ability were estimated for seed yield, head diameter, 1,000-seed weight, plant height and oil content. The results revealed that mean squares for GCA were highly significant for head diameter and significant for yield and oil content. Mean squares for SCA were highly significant for 1,000-seed weight and plant height, while those of yield, head diameter and oil content were non-significant. Components of variance showed that the GCA variance was higher than the SCA variance for yield, head diameter and oil content. These results indicated that additive gene action was more important than non-additive gene action for these traits. Inbred line 5A exhibited the highest GCA effects for yield and oil content, followed by the line 2A. Among all the crosses, 2A´5A showed the greatest positive SCA effects for 1,000-seed weight and oil content. Thus, the two inbred lines (2A and 5A) revealed good potential to be used as parents for hybrid.

QTL analysis of agronomic traits in recombinant inbred lines of sunflower under partial irrigation

The objective of the present research was to map QTLs associated with agronomic traits such as days from sowing to flowering, plant height, yield and leaf-related traits in a population of recombinant inbred lines (RILs) of sunflower (Helianthus annuus). Two field experiments were conducted with well-irrigated and partially irrigated conditions in randomized complete block design with three replications. A map with 304 AFLP and 191 SSR markers with a mean density of 1 marker per 3.7 cM was used to identify QTLs related to the studied traits. The difference among RILs was significant for all studied traits in both conditions. Three to seven QTLs were found for each studied trait in both conditions. The percentage of phenotypic variance (R2) explained by QTLs ranged from 4 to 49%. Three to six QTLs were found for each yield-related trait in both conditions. The most important QTL for grain yield per plant on linkage group 13 (GYP-P-13-1) under partial-irrigated condition controls 49% of phenotypic variance (R2). The most important QTL for 1,000-grain weight (TGW-P-11-1) was identified on linkage group 11. Favorable alleles for this QTL come from RHA266. The major QTL for days from sowing to flowering (DSF-P-14-1) were observed on linkage group 14 and explained 38% of the phenotypic variance. The positive alleles for this QTL come from RHA266. The major QTL for HD (HD-P-13-1) was also identified on linkage group 13 and explained 37% of the phenotypic variance. Both parents (PAC2 and RHA266) contributed to QTLs controlling leaf-related traits in both conditions. Common QTL for leaf area at flowering (LAF-P-12-1, LAF-W-12-1) was detected in linkage group 12. The results emphasise the importance of the role of linkage groups 2, 10 and 13 for studied traits. Genomic regions on the linkage groups 9 and 12 are specific for QTLs of leaf-related traits in sunflower.

Creating new genetic variability in sunflower using induced mutations

SUMMARY The objective of the study was to provide new genetic variability in important agronomic traits that can be exploited for improvement of sunflower production. Seeds of eight sunflower inbred lines from gene collection of Institute of Field and Vegetables, Novi Sad were irradiated with gamma rays (γ) and fast neutrons (Nf) and treated in ethyle-methane-sulphonate (EMS) solution. The manifestation of mutations was mostly expressed in M 2 and M 3 generation. Seven mutants were developed; one early flowering, two short stature and one high, two with higher oil content and one branching. The stable progenies were evaluated in micro-plot tests in M 6 generation for seed yield and other traits in comparison with respective original line. Further studies should be focused on testing new mutant lines in hybrid combinations, as well as the determination of inheritance of mutant traits.

Sunflower inbred lines screening for tolerance to white rot on stalk

Seventy sunflower (Helianthus annuus L) inbred lines were screened for tolerance to white rot on stalk. Plants were inoculated at the budding stage with 4-day old Sclerotinia mycelium grown on PDA medium. Mycelium was placed on the leaf top and covered with tin foil, and the leaf was put into transparent nylon bag in order to maintain high humidity. Spot length on leaf was measured and plant tolerance was determined at the full flowering stage. Obtained results were analyzed by analysis of variance. Tested lines showed significant variability in response to disease. Eleven lines had lesion length less than 50% compared to susceptible control, so they could be considered tolerant and potentially interesting for breeding program.

Development of sunflower hybrids tolerant to tribenuron methyl

Discovery of tribenuron-methyl resistant wild Helianthus annuus L. population (ANN-KAN) created an opportunity for expansion of sunflower herbicide resistance breeding program. The aim of this study was development of sunflower hybrids resistant to tribenuron-methyl. Creation of tribenuron-methyl resistant hybrids would enable the use of a wider palette of herbicides for sunflower, more efficient chemical control of Cirsium arvense and more economically profitable post-emergence control of some annual broad-leaves weeds in sunflower. Original populations SURES-1 and SURES-2 are homozygous for resistance to tribenuron-methyl. F1 generations produced from the crossings are completely resistant to tribenuron-methyl, pointing out to dominant way of inheritance of this trait. Studies on the exact number of genes controlling the resistance are in progress. Tribenuron-methyl resistance was transferred from original populations into a number of female and male inbred lines of cultivated sunflower. These inbred lines could enable creation of a number of hybrids resistant to tribenuron-methyl. Hybrids SUMO-1-PR, SUMO-2- OR and SUMO-3 are resistant to doubled application dose of tribenuron-methyl. Agronomical characteristics of these hybrids are on the level with the leading conventional sunflower hybrids.

Radiosensitivity of sunflower inbred lines to mutagenesis

SUMMARY For much of the past century, mutagenesis has gained popularity in plant genetics research as a means of inducing novel genetic variation. Induced mutations have been applied for the past 40 years to produce mutant cultivars in sunflower by changing plant characteristics that significantly increase plant yield and quality. The present study is focused on generating baseline data to elucidate the role of genotypic differences in the response of sunflower to induced mutagenesis with the aim of expanding the applicability of the use of induced mutant stocks in the genetic improvement of the crop and in its functional genomics. The strategy adopted was to estimate the optimal treatment conditions (doses of mutagens) through relating the extent of damage in seedling progeny to the exposure levels of the initiating propagates to mutagens. Seeds of fifteen elite sunflower genotypes of commonly used as breeding stocks and grown on commercial scales were treated with a range of mutagens: gamma rays (γ rays); fast neutrons and with ethyl-methane-sulphonate (EMS) at different treatment doses. The three mutagenic agents affected seedling height, reducing it with increasing dosage. Based on the mutagen damage on seedling height, the 50% and 30% damage indices (D 50 and D 30 , respectively) were estimated for the 15 sunflower genotypes for the three mutagens. The D 50 (D 30 ) values for the sunflower lines ranged from 120 to 325 Gy (5 to 207 Gy) for gamma irradiation; 9 to 21 Gy (0.1 to 10 Gy) for fast neutrons and 0.69 to 1.55% (0.01 to 0.68%) concentration of EMS.

An analysis of sequence variability in eight genes putatively involved in drought response in sunflower (Helianthus annuus L.)

With the aim to study variability in genes involved in ecological adaptations, we have analysed sequence polymorphisms of eight unique genes putatively involved in drought response by isolation and analysis of allelic sequences in eight inbred lines of sunflower of different origin and phenotypic characters and showing different drought response in terms of leaf relative water content (RWC). First, gene sequences were amplified by PCR on genomic DNA from a highly inbred line and their products were directly sequenced. In the absence of single nucleotide polymorphisms, the gene was considered as unique. Then, the same PCR reaction was performed on genomic DNAs of eight inbred lines to isolate allelic variants to be compared. The eight selected genes encode a dehydrin, a heat shock protein, a non-specific lipid transfer protein, a z-carotene desaturase, a drought-responsive-element-binding protein, a NAC-domain transcription regulator, an auxin-binding protein, and an ABA responsive-C5 protein. Nucleotide diversity per synonymous and non-synonymous sites was calculated for each gene sequence. The π (a)/π (s) ratio range was usually very low, indicating strong purifying selection, though with locus-to-locus differences. As far as non-coding regions, the intron showed a larger variability than the other regions only in the case of the dehydrin gene. In the other genes tested, in which one or more introns occur, variability in the introns was similar or even lower than in the other regions. On the contrary, 3'-UTRs were usually more variable than the coding regions. Linkage disequilibrium in the selected genes decayed on average within 1,000bp, with large variation among genes. A pairwise comparison between genetic distances calculated on the eight genes and the difference in RWC showed a significant correlation in the first phases of drought stress. The results are discussed in relation to the function of analysed genes, i.e. involved in gene regulation and signal transduction, or encoding enzymes and defence proteins.

Studies on pathogenicity in &lt;i&gt;Plasmopara halstedii&lt;/i&gt; (sunflower downy mildew)

Studies on morphological and genetic variabilities as well as alterations in pathogenicity were carried out in Plasmopara halstedii (sunflower downy mildew) isolates of several races. Aggressiveness was analyzed in one sunflower inbred line showing a high level of quantitative resistance. Viability analyses were performed on oval and spheric zoosporangia. Genetic relationships were detected between the pathogen isolates using 12 EST-derived markers. There were significant differences between pathogen isolates for aggressiveness criteria. Based on virulence and aggressiveness reaction for P. halstedii to D3 line of sunflower, there were two groups as more aggressive and less virulent isolates of 100 and 3xx races, and less aggressive and more virulent isolates of 7xx races. The oval zoosporangia for more aggressive isolates of races 100 and 3xx produced more zoospores than the oval ones for less aggressive isolates of races 7xx. The isolates of races 714, 704 and 314 had an intermediary genetic position between the two isolates of races 100 and 710. It is concluded that the relationship between aggressiveness and oval zoosporangia viability may be established in P. halstedii. No correlation was detected between pathogenicity traits and EST genotypes.Key words: aggressiveness; D3 line of sunflower; EST-derived markers; oval zoosporangia; spheric zoosporangia; virulenceDOI: 10.3126/ijls.v4i0.3481International Journal of Life Sciences Vol.4 2010 pp.48-59

Towards reducing genotype specificity in regeneration protocols after somatic hybridization between cultivated sunflower and wildHelianthusspecies

Successful application of somatic hybridization in sunflower breeding is limited by low regeneration percentage from fusion products. The effect of pre-treatment media composition on shoot regeneration from electrofusion products of wild and cultivated sunflower was investigated in this paper, with the aim to determine if there is room for improvement of the existing regeneration protocols in order to make them less genotype-dependent. Protoplasts isolated from hypocotyls of two inbred lines of cultivated sunflower were electrofused with mesophyll protoplasts of H. mollis and H. maximiliani . The regeneration was induced only on calluses that were grown on the media with low auxin and high cytokinin contents during the development of calluses from microcalluses (pre-treatment), with silver nitrate as a medium component. Regeneration from fusion products was induced by treatment with a high concentration of TDZ or 2,4 D for a short period of time.

Can We Enhance Durable Resistance Against Plasmopara Halstedii (Sunflower Downy Mildew)?

Can We Enhance Durable Resistance AgainstPlasmopara Halstedii(Sunflower Downy Mildew)?Sunflower downy mildew caused byPlasmopara halstediiis one of the most potentially important diseases. So far, a complete, major gene resistance (Pl) has been used successfully, but with the appearance of eight races in France since 2000, research on more durable resistance was undertaken. In this study, we present new results concerning the evolution of pathogenicity ofP. halstediiunder conditions of re-enforced infection and differentPlgene selection pressures. Moreover, we imagine the evolution of virulence and aggressiveness ofP. halstediiunder a mixture model of sunflower inbred lines carrying the two types of resistance which may enhance durable resistance against it. Examples of host-parasite interactions including the influence of plant mixture models against pathogens are equally presented to understand how the pathogen develops its pathogenicicity.

Sterol content in sunflower seeds ( Helianthus annuus L.) as affected by genotypes and environmental conditions

Phytosterols play essential roles in many plant cell mechanisms. They are of industrial interest since, as part of the diet, they can reduce low density lipoprotein cholesterol. An increase in plant sterol contents, by improved crop varieties or crop management, could help to answer industrial demands and also to develop environmentally friendly extraction methods. The aim of this study was to evaluate genotypic variability of sterol content in cultivated sunflower and, in particular, effects of sowing date. Results showed large variability among a collection of sixteen sunflower inbred lines and hybrids. Total sterols varied almost twofold between extreme genotypes. A delay of sowing, giving higher temperatures during seed formation, induced a general increase in total sterol concentration by up to 35%, as well as variation in sterol composition according to genotype. These results are considered with an aim of improving sterol content by sunflower breeding programmes.

Identification of sunflower (Helianthus annuus, Asteraceae) hybrids using simple-sequence repeat markers

Hybrid identification of 16 sunflower hybrids was confirmed using simple-sequence repeat methodology. Of 20 specific simple-sequence repeat primers, 18 authenticated the purity of these hybrids; the remaining two specific primer pairs gave ambiguous DNA fragments. The results indicate that simple-sequence repeat analysis for the identification of hybrids derived from the crossing of different inbred sunflower lines can improve the accuracy of selection, save time and reduce cost.