Meiosis and pollen grain viability in Helianthus mollis,Helianthus salicifolius,Helianthus maximiliani and their F1 hybrids with cultivated sunflower

Three diploid perennial sunflower species are useful for variety improvement: Helianthus mollis, because of sessile leaves, H. salicifolius, because of a high oil concentration, and H. maximiliani, a potential source of resistance to Sclerotinia sclerotiorum. The crossability of these species to cultivated sunflower was examined.Hybrids were obtained from eight combinations, with 3–15 F₁ plants per combination. The F₁'s exhibited the dominant phenotype of the wild species. Pollen viability varied between 32.1 and 69.9%. Meiosis was irregular in the F₁ hybrids. At diakinesis, bivalents (62.7–97.9% of meiocytes), univalents (0–31.23%), and multivalents (3.84–7.68%) were detected. At anaphase I, chromosome bridges were detected in 6.77 to 11.44% of meiocytes. Fast chromosomes in metaphase I, and lagging chromosomes in anaphase I and telophase II were evidenced in a high percentage of meiocytes.     

Production of intergeneric hybrids between Brassica and Sinapis species by means of embryo rescue techniques

Reciprocal crosses were made to produce intergeneric hybrids among seven species of Brassica and three species of Sinapis with the aid of embryo rescue techniques. Ovule culture showed better responses in terms of hybrid plant development. Altogether, hybrids from 8 different combinations were obtained from the crosses between B. campestris var. trilocularis × S. turgida, B. campestris var. pekinensis × S. arvensis, S. arvensis × B. campestris var. pekinensis, B. oleracea var. alboglabra × S. alba, B. oleracea var. alboglabra × S. turgida, B. carinata × S. alba, B. carinata × S. arvensis, and B. carinata × S. turgida. Reciprocal crosses yielded no hybrid except in the combination of S. arvensis × B. campestris var. pekinensis. Among them hybrids from 6 combinations were established in the glass house. The hybridity of the plants was confirmed by morphological characters, pollen stainability, chromosome number and by isozyme analysis. Hybrids of four combinations out of six turned out as true hybrids, one as sesquidiploids and the another one as false hybrid plant. This revised version was published online in August 2006 with corrections to the Cover Date.     

Cytogenetic study of Helianthus laevigatus and its F1 and BC1F1 hybrids with cultivated sunflower, Helianthus annuus

Although the wild sunflower species Helianthus laevigatus has not been extensively studied it may be considered for sunflower breeding as a potential source of desirable genes for Sclerotinia stalk rot resistance and high contents of proteins and linoleic acid in the seed. A set of six H. laevigatus populations was crossed to cultivated sun~ower lines and produced nine F₁ (2-14 plants) and 66 BC₁F₁ hybrid combinations (1-13 plants). Male sterility occurred in F₁ and BC₁F₁ hybrid combinations and pollen viability was lower in the progenies than in the parents (51.6-77.2%in F₁ and in F₁ and 4.8-34.0% in BC₁F₁). Meiosis was normal in the H. laevigatus populations It was found that this tetraploid species also occurred in a hexaploid form Numerous irregularities were observed in the meiosis of the F₁ interspecific hybrids During diakinesis, quadrivalents and hexavalents were recorded in addition to bivalents Dislocated chromosomes and chromosome bridges were present in the other phases The chromosome number in F₁ was 68 (tetraploid). Irregularities in chromosome pairing were observed in the interspecific hybrids at BC₁F₁. There were many univalents, and trivalents quadrivalents and hexavalents were also present The chromosome number in the BC₁F₁ generation ranged from 34 to 60. The occurrence of meiotic irregularities in the F₁ and BC₁F₁ interspecific hybrids indicates that H. laevigatus and the cultivated sunflower differ in genome constitution.     

Effect of wild Helianthus cytoplasms on agronomic and oil characteristics of cultivated sunflower (Helianthus annuus L.)

Sunflower (Helianthus annuus L.) productions reliance on a single source of cytoplasmic male sterility, PET1, derived from H. petiolaris Nutt., makes the crop genetically vulnerable. Twenty diverse cytoplasmic substitution lines from annual and perennial wild species were compared with the inbred line HA89 over four environments. Pairwise comparisons of alloplasmic male fertile (MF) with euplasmic MF lines suggested that cytoplasm of perennial H. angustifolius increased lodging while perennial cytoplasms of H. mollis, H. grosseserratus, and H. divaricatus reduced head size and consequently reduced yield and will need to be used with caution. Lines having annual species, cytoplasms had no effects on agronomic traits. Comparisons of CMS HA 89 with 10 CMS lines suggested yield‐reducing cytoplasmic effects of perennial H. maximiliani and annual H. annuus PI 413178 and PI 413024. Cytoplasmic effects on oil percentage and fatty acid composition were minimal and should not be of concern for sunflower breeders. In general, most cytoplasms of wild annual Helianthus species accommodate cultivated nuclear genes without drastic adverse interactions, and are potential sources of cytoplasmic diversity for sunflower breeding.     

Interspecific hybridization inBrassica juncea × Brassica hirta using embryo rescue

Summary Interspecific hybrids have been obtained in an otherwise incompatible cross betweenBrassica juncea × Brassica hirta through the in vitro culture of hybrid ovules and ovaries. The best response was observed from ovules and ovaries cultured 10–15 and 5–7 days after pollination respectively on a basal medium supplemented with indoleacetic acid, kinetin and casein hydrolysate. In some cases the basal cut end of the ovaries proliferated to form callus and shoots. The in vitro-derived hybrid seeds varied in their colour, size and shape, and the F₁ plants in the field showed a large diversity in their morphological traits. The hybrids were sterile, and had an intermediate number of chromosomes (2n=30).     

Interspecific hybridization between Cicer arietinum and C. pinnatifidum

An interspecific hybrid between Cicer arietinum cv. GL 769 and a wild species C. pinnatifidum was obtained after emasculation, pollination and application of growth regulators. Ovules were cultured and embryos were later dissected to obtain hybrid plants. These plants were albinos and morphologically resembled C. pinnatifidum. Shrivelled seeds were also obtained in 2% of the crosses, which on germination gave rise to albino plants. These plants did not survive beyond 20 days. The hybrid nature of these plants was confirmed by esterase isozyme studies. Hybrid shoots obtained from germinating embryos were cultured on modified ML-6 medium with BAP 2 mg/1, IAA 0.5 mg/1, where they turned green after 3–4 weeks. Transmission electron microscopy (TEM) studies on leaf sections from green hybrid shoots showed an improvement in the chloroplast structure, with better organized grana.     

Diversity among sources of cytoplasmic male sterility in sunflower (Helianthus annuus L.)

Summary Ten cytoplasmic male sterile (CMS) sunflower (Helianthus annuus L.) lines were crossed with nine maintainer or male fertility restorer lines in a diallel crossing scheme. Based on fertility restoration of the F₁ generation, CMS lines were divided into four groups. At least two new sources of CMS, CMS PET2 and CMS GIG1, were found to be potentially useful for commercial production of hybrids. Environment had an influence on fertility restoration of one CMS line, CMS MAX1. Effective restoration of male fertility for CMS RIG1, CMS ANN2, and CMS ANN3 was not found.     

Intergeneric hybridization between Brassica species and Crambe abyssinica

A protocol for high frequency callus induction and plant regeneration from sunflower (Helianthus annuus L.) anthers is described. Different variables using Murashige & Skoog (MS) basal medium supplemented with 2.0 mg/l α-naphthaleneacetic acid (NAA) and 1.0 mg/l N₆-benzyladenine (BA) were tested for their ability to enhance the frequency of anther callusing and subsequent embryogenesis. Of these, agar concentration, sucrose concentration, carbohydrate source had significant effect on callusing, while differences due to incubation under dark vs light conditions, cold pretreatment of capitula for 1 to 6 days prior to anther inoculation and genotype on callusing were non-significant. However, all these factors exerted highly significant influence on embryogenesis when calli from the various media were transferred to medium supplemented with 0.1 mg/l NAA and 0.5 mg/l BA. With the procedure developed, callusing as high as 100% and embryo formation at a frequency of 44% was achieved. Although complete embryos were formed the frequency of their conversion to whole plantlets was low (14.3%). Hence, the embryogenic pathway was bypassed to obtain multiple shoots by transferring embryogenic calli with developing embryos to MS medium supplemented with 0.5 mg/l BA. Elongated shoots rooted on half-strength MS medium supplemented with 0.5 mg/l NAA. Cytological analysis of embryogenic callus and somatic embryos revealed haploids at a frequency of 30% while that of rooted plants showed haploid regenerants at a frequency of 8.3%. Nevertheless, the frequency of putative haploid plants could be enhanced through mass multiplication using nodal explants of the regenerants. This revised version was published online in August 2006 with corrections to the Cover Date.     

Interspecific hybridisation of soybeans and perennial Glycine species indigenous to Australia via embryo culture

Summary Five sterile hybrids (2n=59) between a synthetic amphiploid of Glycine tomentella (2n=38) and G. caneseens (2n=40) as female and soybean cultivars Lincoln and Hark as males have been produced by embryo or ovule culture using transplanted endosperm. The hybrid plants are twining perennials like the female parent but possess a number of morphological characters which reflect the presence of the soybean genome. Indophenol oxidase isozymes from leaf extracts also provide good evidence of the hybrid nature of the cultured plants. These hybrids open the way for the exploitation of the diverse germplasm resources of the perennial Glycine species in soybean breeding.     

Cytogenetic studies in hexaploid Helianthus species and their F1 hybrids with cultivated sunflower, H. annum

Hexaploid sunflower species provide potential sources of desirable genes in breeding for resistance to pathogens as well as high protein and oleic acid content. The possibility of using them in sunflower breeding was studied by testing the crossability of hexaploid species to cultivated sunflower, by the analysis of meiosis and pollen viability in the F₁ interspecific hybrids. The hybrids were obtained from 16 combinations, with 1–18 F₁ plants per combination. The F₁ hybrids in general exhibited the dominant phenotype of the wild species. Sterility ranged from 0 to 87.5% per F₁ combination, while pollen viability ranged from 43.0 to 70.2%. Meiosis was almost normal in the species analysed but irregular in the F₁ hybrids. Most of the meiocytes showed predominantly bivalents (86.79–90.22%), but univalents (0.00–4.99%) and the multivalents (8.23–10.09%) occurred as well. At anaphase I, chromosome bridges were detected in 6.88–8.48% of meiocytes. Fast chromosomes in meta-phase I and lagging chromosomes in anaphase I and telophase II were observed in many meiocytes.     

High regeneration potential in vitro of sunflower (Helianthus annuus L.) lines derived from interspecific hybridization

Successful selection of interspecific hybrid progenies with superior ability to regenerate shoots from apical meristems was performed in sunflower which now allows for the development of lines for improved biotechnological applications. Early generations of interspecific hybrids originating from crosses between the two H. annuus CMS lines ‘HA89’ and ‘Baso’, and 9 wild species were screened for their ability to regenerate in vitro. Evaluation of 36 progenies allowed to identify seven progenies from crosses involving H. mollis, H. giganteus, H. strumosus, and H. decapetalus which showed a significantly higher regeneration potential than the commercial hybrid ‘Albena’ regarding the number of shoots per explant. Among these progenies, 47.2 to 62.4% of explants produced shoots with an average of 2.3 to 3.5 shoots per cultured explant. Regeneration in vitro was significantly determined by the genotype. More than half of the investigated interspecific hybrids performed better than the inbred ‘HA89’ demonstrating that the high regeneration potential available in the wild species can be efficiently transferred to cultivated sunflower. The seven progenies with high regeneration potential in vitro were characterised by agronomic performance in the field. Two of the interspecific hybrids derived from H. strumosus and H. decapetalus not only showed a superior regeneration potential but also proved to be competitive to commercial hybrids with regard to important agronomic traits, e.g. fat content and TGW. This revised version was published online in July 2006 with corrections to the Cover Date.     

白菜型油菜黄籽沙逊和芥蓝种间杂交合成甘蓝型油菜研究

人工合成甘蓝型油菜是拓宽油菜遗传育种种质资源的有效途径,也是研究芸薹属物种起源与演化的重要手段之一。为了把黄籽沙逊的黄籽性状同芥蓝的优良特性相结合,本研究利用白菜型油菜黄籽沙逊(Brassica. campestris var. Yellow Sarson Pain, 2n=AA=20)与芥蓝(Brassica albograbra, 2n=CC=18)开展了芸薹属种间杂交研究,在MS+6-BA 1.0 mg/L+NAA 0.1 mg/L+7%蔗糖+活性炭1.0 mg/L的培养基上,通过胚珠培养从授粉7 d的140个胚珠中得到6个远缘杂交胚发育而成的植株,其中1个自然加倍成为双二倍体,随后应用形态学、细胞学和分子标记的方法对双二倍体种间杂种植株进行了鉴定,结果表明杂种植株生长势较强,形态属于中间类型;根尖染色体压片观察显示杂种植株具有父母本的染色体数量之和;SSR分子标记分析表明,杂种植株包含了双亲的遗传信息,由此证明了所获杂种的真实性,这为最后自交分离获得黄籽甘蓝型油菜提供了科学依据。     

Interspecific hybridization in the genus Gossypium through embryo culture

Summary With embryo rescue and culture techniques interspecific hybrids have been obtained amongst diploid cultivated (Gossypium arboreum, G. herbaceum) and wild species (G. stocksii, G. anomalum) of cotton. The early abscission of the young bolls was prevented by repeated application of growth regulators followed by the culture of immature hybrid embryos (15 days after pollination). The best growth and development were obtained on modified Murashige and Skoog's medium with supplements. The plants were stouter when the cultures were first incubated in the dark for 15–25 days, followed by exposure to light. The hybrid plants transferred to soil developed further and matured, and were more or less intermediate between their respective parents.     

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.     

Crossability and embryo rescue enhancement in wide crosses between wheat and three Agropyron species

Summary Soft winter wheat lines were crossed with Agropyron intermedium, A. elongatum and A. trichophorum using pollen from single plants of Agropyron spp. to pollinate wheat spikes. Not only species but also individual plants within varieties of Agropyron species differed in percent seed set with a wheat genotype. In two arrays of crosses between two phenotypically different plants of A. elongatum and nine wheat lines, one Agropyron plant gave higher seed set (overall=27.1%) than the other (overall=3.7%). The differences were significant in seven of the nine cross combinations. Results are consistent with the hypothesis that these two plants differ in their crossability as pollen parents with wheat, and suggest the possibility of occurrence of crossability genes in wheatgrasses. The success rate of hybrid embryo rescue was higher (87.5%) with cold treatment (4°C) than without cold treatment (75.0%) of excised embryos on culture media. Results underscore the significance of genotype of the alien species for crossing with low crossable wheats, and of the physical factors for improving embryo rescue in wide crosses.Contribution No. 11,825, Purdue Agric. Exp. Stn., West Lafayette, Indiana 47907, USA. The research was supported in part by Public Varieties of Indiana.     

Interspecific crosses between Pelargonium×hortorum and P. quinquelobatum using embryo rescue and molecular characterization of hybrids by an endogenous chs probe

Interspecific hybridizations were performed between pelargonium × hortorum (subgenus Ciconium) and a yellow-flowered species. Pelargonium qitinquelobatum(subgenus Eumurpha). Hybrids, using embryo rescue, were obtained only when P. quinquelobatum was used as the female parent. A polymerase chain reaction (PCR) cloned Pelargonium chs gene fragment was used as probe to characterise hybrids hy restriction fragment length polymorphism (RFLP) analysis. Chromatographic flavonoid analysis showed that hybrids could synthesize both flavonol glycosides and anthocyanin in the petals which were pale yellow to pale pink from the proximal to the distal part.     

Facilitation of Wide-Crossing Through Embryo Rescue and Pollen Storage in Interspecific Hybridization of Cultivated Allium species

Up to 70% success has been achieved using a simple technique of in ovulo embryo rescue for generating progeny From crosses between Allium cepa and A. fistulosum and other wild species of Allium. Pollen storage in liquid nitrogen has overcome problems associated with lack of synchrony of flowering between Allium species. The use of pollen tube growth for predicting crossability is brought into question.     

Prefertilization barriers to interspecific hybridization involving Gossypium hirsutum and four diploid wild species

Interspecific hybridization among species of cotton has lead to improvement in productivity, earliness, fibre quality and resistance to pests and diseases. However, wide crosses is often limited by the operation of either pre‐ or/and post‐fertilization barriers. An investigation on pollen tube behaviour of four wild species in the pistils of Gossypium hirsutum was taken up. Pollen germination was normal in crosses involving Gossypium triphyllum and Gossypium armourianum and markedly inhibited in the crosses involving Gossypium davidsonii and Gossypium thurberi. Pollen tubes reached the pistils and fertilization was accomplished within 8 h after pollination (HAP) in control cross. Even though delay in pollen tube was a common phenomenon in all the four crosses successful fertilization was observed in crosses involving G. triphyllum and G. armourianum, as they reached the ovary at 24 HAP. In crosses with G. davidsonii and G. thurberi, pollen tubes failed to reach the ovary even at 24 HAP indicating the presence of strong stylar and ovarian incompatibility. Measures to overcome such barriers to interspecific hybridization in the incompatible crosses are discussed.     

Interspecific hybridization between Vigna unguiculata (L.) Walp. and V. vexillata (L.) A. Rich. through in vitro embryo culture

Immature embryos resulting from the cross V. vexillata × V. unguiculata were cultured on MS medium supplemented with 2,4-D (2 mg/l) and resulted in embryogenic calli. Thirteen hybrid regenerants were obtained via organogenesis by subculturing the calli on MS medium supplemented with BAP (2 mg/l) + adenine sulphate (40 mg/l) + CH (500 mg/l) + cowpea tender pod extract (10%). The interspecific regenerants showed intermediate morphological traits between the parents for leaf shape, pod colour and seed coat colour. The hybrid plants inherited stem, leaf and pod hairiness of the wild species which could serve as a mechanical barrier against viral vectors. Electrophoretic studies of two isozyme systems, peroxidase and esterase, also confirms the hybrid nature of the regenerants as they expressed unique bands of both parents. Cytological study of the meiotic chromosomes revealed high frequency of univalent formation in the hybrids suggesting that the genomes of the parental species are structurally differentiated. The hybrid regenerants exhibited high enzyme activity for three enzymes viz., peroxidase, polyphenol oxidase and phenyl alanine ammonia lyase over the cultivated parent which may be useful in conferring resistance against viral pathogens. This revised version was published online in July 2006 with corrections to the Cover Date.