Saponin polymorphism in the Korean wild soybean (Glycine soja Sieb. and Zucc.)

Seed saponin composition of 3025 wild soybean (Glycine soja Sieb. and Zucc.) accessions collected from nine regions of Korea was analysed by thin‐layer chromatography to determine its polymorphic variation and geographical distribution and find mutants in saponin components. The saponin composition of seed hypocotyls was primarily divided into seven phenotypes, designated as Aa, Ab, AaBc, AbBc, Aa+α, AaBc+α and AbBc+α. The predominant phenotypes were AaBc (55%), Aa (33%), AaBc+α (7.5%) and Aa+α (3.3%). The frequencies of Ab, AbBc and AbBc+α were very low (0.3‐0.5%). Codominant alleles Sg‐1^a and Sg‐1^b and dominant allele Sg‐4 occupied 98.6, 1.1 and 63.3%, respectively. Alleles Sg‐3 and Sg‐5 were found to be dominant in all the analysed accessions except the mutants. Three accessions were discovered as mutants via LC‐PDA/MS/MS. The accession CWS0115 did not produce saponin Aa and Ax, CWS2133 did not produce saponin Aa and Ab and CWS5095 did not produce any group A saponins. These newly determined mutants might be utilized in producing a new soybean variety with good taste as well as in biosynthetic studies.     

Genetic analysis of variations in the sugar chain composition at the C-3 position of soybean seed saponins

Saponins are sterols or triterpene glycosides that are widely distributed in plants. The biosynthesis of soybean saponins is thought to involve many kinds of glycosyltransferases, which is reflected in their structural diversity. Here, we performed linkage analyses of the Sg-3 and Sg-4 loci, which may control the sugar chain composition at the C-3 sugar moieties of the soybean saponin aglycones soyasapogenols A and B. The Sg-3 locus, which controls the production of group A saponin Af, was mapped to chromosome (Chr-) 10. The Sg-4 locus, which controls the production of DDMP saponin βa, was mapped to Chr-1. To elucidate the preference of sugar chain formation at the C-3 and C-22 positions, we analyzed the F₂ population derived from a cross between a mutant variety, Kinusayaka (sg-1⁰), for the sugar chain structure at C-22 position, and Mikuriya-ao (sg-3), with respect to the segregation of the composition of the group A saponins, and found that the formation of these sugar chains was independently regulated. Furthermore, a novel saponin, predicted to be A0-γg, 3-O-[β-d-galactopyranosyl (1→2)-β-d-glucuronopyranosyl]-22-O-α-l-arabinopyranosyl-soyasapogenol A, appeared in the hypocotyl of F₂ individuals with genotype sg-1⁰/sg-1⁰ sg-3/sg-3.     

Identification of a novel variant lacking group A soyasaponin in a Chinese wild soybean (Glycine soja Sieb. & Zucc.): implications for breeding significance

Group A saponins are the principal factors underlying the undesirable bitter and astringent tastes in soybean food products. Therefore, the genetic reduction or elimination of the undesirable tastes is of great significance in soybean taste breeding. Using thin‐layer chromatography and liquid chromatography–mass spectrometry, we identified a novel mutant lacking group A saponins in a wild soybean (Glycine soja Sieb. & Zucc.), reflecting the failed arabinose binding of arabinosyltransferase to the C‐22 hydroxyl group, consequently leading to a lack of group A saponins and the concomitance expression of two new components A‐αg and KA‐αg in seeds. The C‐3 glycosylated soyasapogenol A (SS‐A), A‐αg, is a steady precursor compound in biosynthesis of Aa and Ab types in the seeds of soybean plants and exhibits stable inheritance. In this study, we observed a critical step necessary for arabinose binding to the C‐22 hydroxyl group in the biosynthetic pathway of group A saponins and identified an important germplasm accession for the genetic improvement in the tastes of soybean milk and processed soybean foods.     

Characterization of an EMS-induced soybean mutant with an increased content of Af saponin and a new component Ab-δ in the seed hypocotyl

Saponins are one of the components present in the soybean seeds that have various functional properties. The chemical structures and concentration of soyasaponins affect the taste of the processed soyfood, thereby limiting its industrial applications. Therefore, it is important to understand saponin biosynthesis to explore natural and artificial variation in the saponin components, which can be modified to suit its application. The objective of the present study was to identify and characterize an EMS-induced soybean mutant with an altered saponin composition from a pool of 892 M4 lines. The mutant PE1905 showed an increased content of saponin Af (336.0%). The content of saponin Ab, DDMP-αg, and DDMP-βg was decreased in the mutant PE1905 by 89.3, 24.8, and 63.1%, respectively compared to the wild-type Pungsannamul. Additionally, four new components were detected in the mutant PE1905 that were absent in the wild type. Of these, the compound 4 (designated as Ab-δ) had the highest concentration, and therefore it was further characterized by HPLC and LC-PDA/MS/MS analysis to know the chemical structure, and molecular weight and formula. Considering these details, along with the alterations in the saponin Af and Ab concentrations, it was presumed that the Ab-δ acts as a precursor for the synthesis of saponin Af and Ab. Thus, we predicted a biosynthetic pathway from the Ab-δ to Ab saponin. The inheritance analysis showed that the concentration of saponin Ab-δ is controlled by a single recessive gene in the mutant PE1905. The results from the present study would be helpful in understanding the mechanisms behind altered seed saponin composition in soybeans.     

Stability of elevated α-linolenic acid derived from wild soybean (Glycine soja Sieb. & Zucc.) across environments

Previous studies reported that omega-3 fatty acid and α-linolenic acid are important compounds that prevent cardiovascular disease and cancer in humans. Soybean [Glycine max (L.) Merr.] oil typically contains ~8 % α-linolenic acid (ALA). Elevated (~15 %) ALA content in seed oil is a trait of wild soybeans (G. soja Sieb. and Zucc.). Decreasing the ratio of linoleic acid (LA) to ALA to 4:1 or lower (compared to the ratio of 6 or 7:1 found in commercial soybean seed) should have health benefits for humans. This study was conducted to determine the environmental stability of elevated ALA acid recombinant inbred lines (RILs) derived from a cross of PI 483463 (wild soybean with 15 % ALA) and Hutcheson (cultivar with 9 % ALA). The fatty acid profile analysis from nine environments showed that the content of ALA for the RILs 156, 159 and 166 ranged from 10.7 to 15.7, 14.0 to 15.8, and 14.8 to 15.8, and averaged 13.9, 14.9, and 15.2 % respectively. The contents of ALA from these RILs and the wild soybean parent showed consistently higher than cultivated check soybeans. Two of the three RILs with elevated ALA content and ratios of <4:1 LA to ALA were as stable in ALA content as the high and low linolenic acid parents across growing environments. This indicates that lines with elevated ALA content developed from wild soybean PI 483463 are stable in ALA across environments and would be useful in improving soybeans with lower LA to ALA ratios.     

Genome-wide detection of genetic loci associated with soybean aphid resistance in soybean germplasm PI 603712

Soybean aphid (Aphis glycines Matsumura) has become one of the major pests of soybean [Glycine max (L.) Merr.] in North America since 2000. At least four biotypes of soybean aphid have been confirmed in the United States. Genetic characterization of new sources of soybean aphid resistance will facilitate the expansion of soybean gene pool for soybean aphid resistance and thus will help to develop soybean aphid resistant cultivars. To characterize the genetic basis of soybean aphid resistance in PI 603712, a newly identified resistant germplasm line, 142 F₂ plants derived from the cross ‘Roberts’ × PI 603712 and their parents were evaluated for soybean aphid resistance in the greenhouse, and were genotyped with BARCSoySNP6K Illumina Infinium II BeadChip. A genome-wide molecular linkage map was constructed with 1495 polymorphic SNP markers. QTL analysis revealed that PI 603712 possessed two major loci associated with soybean aphid resistance, located on chromosome 7 and 16, respectively. The locus on chromosome 7 was dominantly expressed and positioned about one Mega-base-pair distant from the previously identified resistance locus Rag1. The locus on chromosome 16 was positioned near the previously identified resistance locus Rag3 and expressed partially dominance or additive effect. Interestingly, two minor loci were also detected on chromosomes 13 and 17 but the alleles from PI 603712 decreased the resistance. In developing soybean aphid resistant cultivars through marker-assisted selection, an appropriate combination of resistance loci should be selected when PI 603712 is used as a donor parent of resistance.     

Production and characterization of interspecific hybrids between Asparagus kiusianus Makino and A. officinalis L.

Interspecific hybridization is a useful technique to introduce characteristics from wild species into crops. Garden asparagus (Asparagus officinalis, 2n = 2x = 20) is an economically important vegetable that is native to the Mediterranean region but widely cultivated in many countries. The genus Asparagus is comprised of over 100 species. Asparagus kiusianus (2n = 2x = 20) is a wild asparagus species endemic to Japan. This species occurs on the coast and is likely to be a salt-resistant species. Although the geographic distribution of these two species is not close, molecular phylogenetic analysis demonstrated that these two species are closely related. In this study, a reciprocal cross between A. officinalis and A. kiusianus was carried out by hand pollination, and progeny were obtained from both crossings. These progeny exhibited a morphologically intermediate phenotype in terms of flower shape, and restriction fragment length polymorphism analysis confirmed that these were indeed interspecific hybrids. The interspecific hybrids were fertile, and backcross progeny with garden asparagus was also generated. These interspecific hybrids are expected to supply novel traits to garden asparagus.     

Mapping and use of QTLs controlling pod dehiscence in soybean

While the cultivated soybean, Glycine max (L.) Merr., is more recalcitrant to pod dehiscence (shattering-resistant) than wild soybean, Glycine soja Sieb. & Zucc., there is also significant genetic variation in shattering resistance among cultivated soybean cultivars. To reveal the genetic basis and develop DNA markers for pod dehiscence, several research groups have conducted quantitative trait locus (QTL) analysis using segregated populations derived from crosses between G. max accessions or between a G. max and G. soja accession. In the populations of G. max, a major QTL was repeatedly identified near SSR marker Sat_366 on linkage group J (chromosome 16). Minor QTLs were also detected in several studies, although less commonality was found for the magnitudes of effect and location. In G. max × G. soja populations, only QTLs with a relatively small effect were detected. The major QTL found in G. max was further fine-mapped, leading to the development of specific markers for the shattering resistance allele at this locus. The markers were used in a breeding program, resulting in the production of near-isogenic lines with shattering resistance and genetic backgrounds of Japanese elite cultivars. The markers and lines developed will hopefully contribute to the rapid production of a variety of shattering-resistant soybean cultivars.     

Genetic characterization of group A acetylsaponin‐deficient mutants from wild soybean (Glycine soja Sieb. and Zucc.)

Group A acetylsaponins are the main causative components for bitter and astringent tastes of soybean (Glycine max). In this study, we examined the genetic nature of the absence of group A acetylsaponins in 12 Korean wild soybean (Glycine soja) accessions. In all 12 accessions, the coding region (1431‐bp) of Sg‐1 locus was identical with Sg‐1^a, which adds the xylose sugar moiety at the terminal position of the C‐22 sugar chain of SS‐A, except one nucleotide (G→A change) at +948th position. This point mutation results in change of one amino acid from tryptophan (TGG) to stop codon (TGA). We observed that the mutated Sg‐1 was controlled by a single recessive gene (sg‐1^0‐a1). This gene was mapped between BARCSOYSSR_07_1561 and BARCSOYSSR_07_1598 on soybean chromosome 7. Our study demonstrated that the mutated Sg‐1 gene in Korean wild soybeans is genetically different from those identified in Japanese soybean cultivar ‘Kinusayaka’ and wild soybean JP‐36121. We believe that the new Sg‐1 mutants can also be utilized to produce a new soybean variety without bitter and astringent properties.     

Flavan-3-ols and procyanidins in grape seeds: biodiversity and relationships among wild and cultivated vines

Flavan-3-ol monomers and polymers composition of seeds from wild (17) and autochthonous (8) Vitis vinifera grapes growing in northern Tunisia were evaluated. Wild grape seeds were spherical with a small beak and relatively a high seed/berry ratio (~ 18.1%w/w). Local cultivars developed pyriform-shaped seeds with a well-developed beak representing on average 2.2% of total weight of the berry. Flavanol concentrations ranged between 40.9 and 67.5 mg/g FW in seeds from wild accessions and between 48.9 and 96.7 mg/g FW in seeds from cultivated grapes. Differences between accessions were highly significant (p < 0.01) and seeds from cultivar ‘Boukhasla’ showed the highest polyphenols content. Among flavan-3-ol monomers, (+)-catechin was predominant for all ecotypes and generally their abundance was: (+)-catechin (Cat) > (?)-epicatechin (Ec) > (?)-epicatechin-3-O-gallate (EcG). The Cat/Ec ratio was approximately 1.7 for wild grapes while it was about 2.5 for cultivated grapes. Procyanidins in wild seeds differed from cultivated ones by a lower mDP and higher proportions of galloylated derivatives, likely to affect fruit bitterness and astringency. (?)-epicatechin was the main extension subunit in grape seed procyanidins, reaching on average 52% in wild and 58% in cultivated seeds. Hierarchical cluster analysis based on seeds morphometry and procyanidin profile indicated close proximity between some wild and cultivated grapes suggesting that some cultivars derived from ancestral events of local domestication or cross hybridization with native wild plants.     

Breeding of tetra null soybean (Glycine max) for lipoxygenase,kunitz trypsin inhibitor,lectin, and 7S α' subunit proteins

Soybean seed includes various bioactive substances. Also, they contain a variety of antinutritional factors including lipoxygenase, Kunitz trypsin inhibitor (KTI), lectin, and 7S α' subunit proteins. The genetic removal of these proteins will improve the nutritional value of soybean seed. The objective of this research was to breed new soybean with tetra recessive alleles (lox1lox2lox3/lox1lox2lox3‐ti/ti‐le/le‐cgy1/cgy1) for lipoxygenase, KTI, lectin, and 7S α' subunit proteins. Seven parents were used to breed tetra null strain. SDS‐PAGE and Western blot analysis were used to determine the presence or absence of lipoxygenase, 7S α' subunit, KTI, and lectin proteins in mature seed. Tetra null soybean line has a purple flower, determinate growth habit, tan pod, and yellow seed coat colour. Stem height of the breeding line was 62.3 cm. The 100‐seed weight of the breeding line was 27.1 g and yield (t/ha) was 2.84. This is the first soybean strain with lox1lox2lox3/lox1lox2lox3‐ti/ti‐le/le‐cgy1/cgy1 genotype (absence of lipoxygenase, KTI, lectin, and 7S α' subunit proteins).     

Grain yield and grain protein percentage of common wheat lines with wild emmer chromosome-arm substitutions

Wild emmer wheat, Triticum turgidum subsp. dicoccoides, (2n = 4× = 28; genome BBAA), the progenitor of domesticated wheat, is genetically closely related to durum and common wheat. This wild taxon has characteristics that would be valuable if transferred to domesticated wheat. A series of chromosome-arm substitution lines (CASLs) of wild emmer wheat were produced in the background of an Israeli common wheat cultivar. These CASLs were evaluated in a pot experiment and in field trials in Israel and California for their grain yield (GY) and its components and for grain protein percentage. In addition, the extent of genetic interactions (epistatic effects) between “wild” and “domesticated” alleles, within and between homoeologous groups 1 and 7 as expressed in grain and protein yields and other quantitative traits, were determined. The research has shown that wild emmer wheat harbors genetic variability for quantitative traits and that the “wild” genes interact among themselves in a non-additive way in the common genetic background. Several chromosome arms improve GY and protein percentage in common wheat, but their effects will be presumably enhanced when combination of genes from several “wild” arms are integrated into a single “domesticated” genotype. Hence, the interaction between these genes and those in the recipient common wheat must be accounted for when higher yield or protein content is desired. The results of this study indicate on the potential of this material for breeding and genetic analysis, and support the idea of pyramiding genes from a wild species.     

陕西省野生大豆种质资源的SSR遗传多样性研究

为了研究陕西地区野生大豆的遗传多样性特点,利用SSR分子标记分析了陕西省6个野生大豆(Glycine soja)天然种群和1个栽培大豆(Glycine max)种群的遗传结构与遗传多样性。结果显示：13个位点共检测出113个等位基因,平均每个位点的等位基因数(A)为8.69个,等位基因数目范围为4~13个,有效等位基因数(N_e)范围为2.135(Satt590)~9.385 (Satt487),平均有效等因基因数为5.623;观察杂合度(H_o)变化范围为0.033~0.121,平均为0.080;预期杂合度(H_e)的变化范围为0.312~0.658,平均为0.482;种群平均Shannon遗传多样性指数(I)为0.657;野生大豆种群基因多样度比率(F_ST)为0.465。该研究显示,陕西省野生大豆具有较高水平的遗传多样性,野生大豆的遗传多样性普遍高于栽培大豆;随着海拔的不断升高,野生大豆遗传多样性变低;陕西中部、南部的野生大豆种质资源丰富、种群具有较高的遗传多样性,推测该区域为陕西省野生大豆的遗传多样性中心。     

Genetic studies on saline and sodic tolerances in soybean

Salt-affected soils are generally classified into two main categories: saline and sodic (alkaline). Developing and using soybean (Glycine max (L.) Merr) cultivars with high salt tolerance is an effective way of maintaining sustainable production in areas where soybean growth is threatened by salt stress. Early classical genetics studies revealed that saline tolerance was conditioned by a single dominant gene. Recently, a series of studies consistently revealed a major quantitative trait locus (QTL) for saline tolerance located on linkage group N (chromosome 3) around the SSR markers Satt255 and Sat_091; other minor QTLs were also reported. In the case of sodic tolerance, most studies focused on iron deficiency caused by a high soil pH, and several QTLs associated with iron deficiency were identified. A wild soybean (Glycine soja Sieb. & Zucc.) accession with high sodic tolerance was recently identified, and a significant QTL for sodic tolerance was detected on linkage group D2 (chromosome 17). These studies demonstrated that saline and sodic tolerances were controlled by different genes in soybean. DNA markers closely associated with these QTLs can be used for marker-assisted selection to pyramid tolerance genes in soybean for both saline and sodic stresses.     

Evaluation of important seed and sprout traits as potential selection criteria in breeding varieties for sprout soybeans

Soybean sprouts, a traditional vegetable in Asia, are gaining popularity in the United States. Soybean sprout demand has been supplied by natto (a Japanese soyfood) cultivars that share some seed characteristics with sprout cultivars. However, natto seeds do not meet all requirements of sprouts and are rejected by sprout manufacturers. The objectives of this study were to evaluate important seed and sprout traits as potential selection criteria in breeding sprout soybeans and to study the storage effect on soybean sprout quality. Almost all genotypes produced thicker and longer hypocotyls and higher fresh-sprouts than ‘MFS-561’, a commercial soybean sprout variety. Hypocotyl length ranged from 13.8 to 16.2 cm. Four fungi genera Bipolaris sp., Cercospora sp., Botrytis sp. and Caethomium sp. were isolated from seeds. Cracked cotyledons and abnormal seedlings were the two main constraints affecting soybean sprout quality. Correlation coefficients among all traits indicated that percentage and weight of high- and average-quality sprouts would determine sprout yield. Acceptable yield and several traits were recommended to be used simultaneously while breeding superior sprout soybean cultivars. Good sprout varieties should produce high-quality sprouts >48%, average-quality sprouts <38%, low-quality sprouts <14%, sprout yield >5.7 g/g seed, hypocotyl thickness >1.6 cm and hypocotyl length >13 cm. One-year seed storage at room temperature reduced sprout quality. V09-3876 and V12-1939 had superior seed and sprout traits and are promising lines for further evaluation for sprout production. Seed storage over time affects seed germination and seedling vigor, and fungi on seed can cause reduced seed quality.     

Influence of the maize Sugary1 locus on genetics of economically important traits

Knowledge of genetic control of mutant viability is of great importance in maize breeding, particularly for mutants with deleterious effects. Little is known about the genetics of the viability of mutants and no previous report has been published concerning the genetic effects of the mutant sugary1 on agronomic traits. Our objective was to study the effects of the sweetcorn mutant sugary1 (su1) on the genetic effects of agronomic traits in two wild type corn backgrounds. Estimated genetic effects of agronomic traits in Su1 versus su1 plants were monitored through four successive selfpollination cycles in two separated mean generation designs. The first involved two Corn Belt inbred lines A619 and A632, while unrelated inbred lines EP42 and A631 were used for the second design. Parents, F₁s, F₂s, and backcrosses were crossed to the su1 inbred P39 as the donor of su1 and the 12 crosses were successively selfpollinated for 4 years. For each cross, Su1 and su1 seeds were separated and evaluated in 10 × 10 triple lattice designs. The Su1 plants showed higher performance than the su1 plants in almost all traits. The estimates of genetic effects of Su1 versus su1 plants were strongly affected by the su1 × wild type corn interactions. The introgression of su1 in wild type corn strongly affects the genetic effects of flowering time and, to a lesser stent, that of other plant or ear traits. Mutant viability is regulated by additive, dominance and digenic epistatic effects and the importance of those genetic effects depends on the genetic background and environmental conditions.     

Intergeneric hybrid between Chrysanthemum × morifolium and Artemisia japonica achieved via embryo rescue shows salt tolerance

Using embryo rescue, we generated an intergeneric hybrid between Chrysanthemum × morifolium ‘Maoyan’ and Artemisia japonica Thunb. Cytological tests confirmed that regenerated plantlets were all genuine hybrids possessing 45 chromosomes, with 27 chromosomes inherited from C. × morifolium (2n = 6x = 54) and the other 18 derived from A. japonica (2n = 4x = 36). Hybrid plant flowered normally. The shape and color of the hybrid flowers and leaves resembled those of chrysanthemum, while leaf width, leaf length, plant height, and inflorescence diameter were intermediate between those of the parents. Hybrid plant had higher levels of chlorophyll and free proline, and lower concentrations of malondialdehyde and Na⁺, than the maternal parent (C. × morifolium), and these levels were correlated with the hybrid’s enhanced salt tolerance. These results clearly demonstrate that intergeneric hybridization is an effective method of cultivar improvement in chrysanthemum.     

Genetic mapping of quantitative trait loci conditioning salt tolerance in wild soybean (Glycine soja) PI 483463

Salt tolerance in soybean [Glycine max (L.) Merr.] is controlled by major quantitative trait loci (QTL) or single gene(s). Among soybean germplasm, wild soybean plant introduction PI 483463 was reported to have a single dominant gene for salt tolerance. The objective of this study was to genetically map the QTL in a recombinant inbred line (RIL) population derived from a cross between PI 483463 and Hutcheson. Simple sequence repeat (SSR) markers and universal soybean single nucleotide polymorphism (SNP) panel (the USLP 1.0) were utilized for molecular genotyping. The RILs were phenotyped in two independent tests in a greenhouse using a 1–5 scale visual rating method. The results showed that the salt tolerant QTL in PI 483463 was mapped to chromosome 3 in a genomic region between the Satt255 and BARC-038333-10036 markers. The favorable allele inherited from PI 483463 conferred tolerance to salinity and had an additive effect on reducing leaf scorch. A subset of 66 iso-lines was developed from the F₃ families of the same cross and was used for genetic confirmation of the QTL. The integration of recombination events and the salt reaction data indicate that the QTL is located in the region of approximately a 658 kb segment between SSR03_1335 at nucleotide 40,505,992 and SSR03_1359 at nucleotide 41,164,735 on chromosome 3. This narrow region can facilitate further genomic research for salt tolerance in soybean including cloning salt tolerance genes.     

Development of a scoring scale for powdery mildew (Golovinomyces cichoracearum (DC.) V.P. Heluta) disease and identification of resistance sources in cultivated and wild sunflowers

Powdery mildew incited by Golovinomyces cichoracearum has become a serious problem on sunflower in India during the past 2–3 years. Genetic resistance in the released cultivars and the parental lines of hybrids is rather limited. Hence, screening of about 420 accessions comprising of wild Helianthus species, interspecific derivatives, core germplasm, inbred lines and few exotic accessions was done under natural field conditions for 2 years and further confirmed by screening under artificial inoculation conditions. PCR analysis using primers specific to powdery mildew causing genera gave a 391 bp band which confirmed the pathogen as G. cichoracearum. Seven different screening methods were tested which induced infection, but dusting of spores on to the healthy leaves proved to be convenient and more effective method of infection. Based on the differential response of the accessions derived from diverse genetic backgrounds, a scale for obtaining reliable estimates of the disease has been devised. Among different cultivar germplasm accessions, the disease severity index (DSI) ranged from 15 to 100 and area under disease progression curve (AUDPC) ranged from 95 to 648. Among the four groups of cultivated sunflower accessions tested, DSI and AUDPC was in the order of exotic lines < interspecific derivatives < inbred lines < core germplasm. Reliable sources of resistance to the pathogen were identified in four annual wild species (H. argophyllus, H. agrestis, H. debilis, H. praecox), six perennials (H. angustifolius, H. atrorubens, H. rigidus, H. salicifolius, H. pauciflorus and H. resinosus), two interspecific derivatives (HIR-1734-2, RES-834-3) and two exotic lines (PI 642072, EC-537925).