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.     

Development and characterization of interspecific hybrids of Trifolium alexandrinum×T. apertum using embryo rescue

This is the first report on the development of interspecific hybrids between Trifolium alexandrinum and T. apertum using embryo rescue and characterization of F₁ plants. T. apertum was used as the male parent and T. alexandrinum as the female parent. Development of interspecific hybrids under natural conditions is not successful and so embryo rescue was attempted. Of the several combinations tried, pollination 2 days after emasculation and embryos rescued 11 days after pollination was found to be the best. For embryo culture, EC3 medium consisting of MS basal supplemented with 2.3 μM kinetin and 3% sucrose was used. Germinated embryos were transferred to LSP3 medium 25 days after inoculation wherein most of the cultures showed multiple shoots that were split and subcultured on RL1 medium for rooting. After hardening, about 75% of hybrids were successfully transferred to the field. The hybrids, in general, showed morphological traits intermediate between the two parents; however, a few hybrids showed better growth than either parent. Some F₁ plants were almost 3 weeks later in flowering than the female parent. Pollen fertility among these plants ranged from 78 to approximately 100%. Chromosomal associations at diakinesis and isozyme banding patterns for acid phosphatase (ACP), superoxide dismutase (SOD) and peroxidase also confirmed the hybrid nature of the plants.     

Interspecific hybridization between sunflower and wild perennial Helianthus species via embryo rescue

Interspecific crosses were made between the cultivated sunflower inbred line HA89 and accessions of five wild perennial Helianthus species (H. giganteus L., H. laevigatus T. & G., H. resinosus Small, H. pauciflorus Nutt. and H. decapetalus L.) resistant to broomrape ( Orobanche cernua Loefl.). Using the genetic male-sterile isogenic version of that line as female, successful reciprocal crosses were also obtained with all these wild species except with H. decapetalus. Five-day-old hybrid embryos were excised and cultured in vitro. In all cases, few mature plants were obtained from embryos in early developmental stages (early heart and globular) but up to 28% mature plants were obtained from embryos in later stages of development. Very immature embryos were difficult to excise without damage. Hybrid embryos and mature plants were obtained from all five wild species. Interspecific hybrid embryos from different wild species showed distinct developmental potentials, the proportion of hybrid embryos in different developmental stages varying among species. Differences in the proportion of hybrid embryos at the several developmental stages were also obtained for the reciprocal crosses (cultivated × wild), which showed higher proportion of fully developed embryos. Hybrids involving H. giganteus and cultivated sunflower were difficult to obtain without the use of embryo culture. Embryo culture proved to be an useful tool to overcome post-zygotic hybrid incompatibility in different Helianthus spp., and facilitated interspecific transfer of resistance to O. cernua. This revised version was published online in July 2006 with corrections to the Cover Date.     

Interspecific compatibility barriers,development of interspecific hybrids through embryo rescue and lineage of Trifolium alexandrinum (Egyptian clover)—important tropical forage legume

Genetic improvement of Trifolium alexandrinum L. is hampered because of narrow genetic base and lack of interspecies compatibility information. The study was envisaged to understand the interspecies incompatibility and the affinity between T. alexandrinum and 22 species of the genus representing different sections and to develop interspecific crosses. Development of interspecific hybrids with T. resupinatum, T. lappaceum, T. subterraneum, T. vesiculosum and T. echinatum, through embryo rescue, revealed their affinity with T. alexandrinum. Failure of pollen germination or restricted pollen tube growth showed its distant relatedness with T. clypeatum, T. balansae, T. purpureum, T. leucanthum, T. hirtum, T. pilulare and T. hybridum. Pollen tubes reached up to ovule among the crosses with T. alpestre, T. repens and T. nigrescens, whereas it travelled up to ovary among the crosses with T. angustifolium, T. grandiflorum and T. dasyurum indicating need of manipulation to overcome barrier. Thus, incompatibility of T. alexandrinum with many Trifolium species could be overcome through embryo rescue with intensive crossing. Egyptian clover ecotype ‘Fahli’ was more compatible with other species than ‘Mescavi’.     

火龙果加工技术

火龙果也称红龙果,系仙人掌科量天尺(三角柱)属的仙人掌果实,形似大芒果,颜色桃红,清甜可口,富含维生素及钙、磷、铁等矿物质,为最佳的天然维他命来源.是一种低热量、低脂肪、高纤维素的水果.以下介绍以火龙果为主料的几种加工产品的制作技术.     

Development of black rot resistant interspecific hybrids between Brassica oleracea L. cultivars and Brassica accession A 19182, using embryo rescue

Black rot is a bacterial disease of Brassica oleracea caused by Xanthomonas campestris pv. campestris. Resistance to the major black rot races 1 or 4 has been identified in related Brassica species including B. carinata and B. napus. In this study, two B. juncea accessions (A 19182 and A 19183) that are resistant to races 1 and 4 of Xcc were used as maternal and paternal parents to generate interspecific hybrids with B. oleracea cultivars. Interspecific hybrids were recovered using the embryo rescue technique and confirmed through inheritance of paternal molecular markers. Twenty-six interspecific hybrid plants were obtained between A 19182 and B. oleracea cultivars, but no interspecific hybrids were obtained using A 19183. Although interspecific hybrid plants were male sterile, they were used successfully as maternal parents to generate backcross plants using embryo rescue. All hybrid and BC₁ plants were resistant to black rot races 1 and 4.     

3种火龙果种子萌发及其幼苗形态的比较研究

红皮白肉、红皮红肉及黄皮白肉火龙果均具有较高的经济价值.对其种子萌发及幼苗形态的差异进行分析,将有助于3种火龙果进行高效的育种和育苗.本实验以上述3种火龙果种子为试验材料,进行火龙果种子萌发及幼苗形态间差异的研究.结果表明:在相同处理条件下,红皮白肉和红皮红肉火龙果种子的发芽率显著高于黄皮白肉火龙果,但在发芽后第14天,它们的幼苗鲜重均显著低于黄皮白肉火龙果.在幼苗移栽后第60天,黄皮白肉火龙果的幼苗鲜重和幼苗长度均显著高于红皮白肉和红皮红肉火龙果,但幼苗根系总长在3种火龙果之间差异不显著.     

Production of two mandarin × trifoliate orange hybrid populations via embryo rescue with verification by SSR analysis

Intergeneric sexual hybridizations were conducted between two genera of Rutaceae, with Satsuma mandarin (Citrus unshiu Marc) and Red tangerine (C. reticulata Blanco) as maternal parents, and Poncirus trifoliata (L.) Raf as the paternal parent, in an effort to generate hybrid populations for both molecular mapping and rootstock breeding. Embryo rescue is important for citrus sexual breeding because polyembryony can interfere with hybrid embryo recovery. Immature embryos of 80, 85, 90 days after pollination (DAP) from the Satsuma mandarin (S) × trifoliate orange (P) cross, 80 and 85 DAP from the Red tangerine (R) × trifoliate orange (P) cross, were cultured on MG1.0 medium consisting of MT basal medium supplemented with 1.0 mg l⁻¹ GA₃ and 4% sucrose. The results showed that 80 DAP was the optimal time for embryo rescue of the tested crosses, as evidenced by embryos at this stage exhibiting high germination rates, 37.3% for S × P and 51.3% for R × P. Among the eight tested media, MT medium supplemented with 0.5 mg l⁻¹ GA₃ was the best one for Satsuma mandarin, and MT plus 1.0 mg l⁻¹ GA₃ for Red tangerine. A total of 85 plants were obtained from S × P, and 340 from R × P. Out of them, 44 progenies from S × P and 111 from R × P were sufficiently confirmed to be hybrids by morphological characterization and SSR analysis. In addition, two hybrid callus lines were obtained from S × P and R × P respectively.     

需光性火龙果种子萌发特性的分析

用种子繁殖火龙果幼苗是非常有价值的育苗方式.本实验深入分析了火龙果种子萌发对温度、光照、赤霉素以及硝酸钾等因子的响应特性.结果表明:细小的火龙果种子在黑暗中不能萌发;播种深度较深时,出苗率急剧下降,甚至在8 cm深时不出苗.在黑暗中,即用赤霉素溶液和硝酸钾溶液预处理和培养,火龙果种子也未能萌发.在光照下,成熟的火龙果种子在8~30℃的范围内可以顺利萌发(100%),其中25℃/17℃下的萌发速率最快.由此可知,火龙果种子是需光性的.在实践中,可在光下育苗;待种子萌发后,再培土养苗是行之有效的方法.     

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.     

Embryo rescue and plant regeneration following interspecific crosses in the genus <Emphasis Type="Italic">Hylocereus</Emphasis> (Cactaceae)

The aim of this study was to develop an efficient methodology to rescue embryos following interspecific crosses in the genus Hylocereus. Crosses between the diploids Hylocereus polyrhizus and H. undatus in both directions were performed. Fertilized ovules carrying embryos at very early pro-embryonic stages were excised from ovaries 5 days after pollination (DAP) and placed on half-strength basal MS medium containing 680 μM glutamine, 0.55 μM α-naphthaleneacetic acid (NAA), 0.45 μM thidiazuron (TDZ) and various concentrations of sucrose. After 30 days in culture, ovules were isolated from the surrounding tissue and transferred to the same fresh medium. Significant differences were found between the main effects (cross and sucrose concentration) in ovule response, i.e., increased ovule size and callus formation. The best responses were obtained in the cross: H. polyrhizus × H. undatus; and sucrose concentration of 0.09 M. In terms of embryo conversion, polyembryony and number of regenerated plants, the highest responses were observed on the culture medium supplemented with 0.17 M sucrose in both interspecific crosses. All tested plants were found to be diploid by flow cytometric analyses. Fluorescent amplified—fragment length polymorphism (fAFLP) confirmed the hybrid origin of the regenerated plants. This study reports on the success of a three-step embryo rescue procedure for Hylocereus species. The procedure developed here provides the means for producing plants from very-early embryo stage, thus expanding the prospects for vine-cactus breeding programs.     

The transfer of triazine resistance from Brassica napus L. to B. oleracea L. I. Production of F1 hybrids through embryo rescue

Summary Triazine resistant Brassica napus ssp. oleifera and ssp. rapifera were hybridized to cultivars of B. oleracea ssp. italica, ssp. botrytis, ssp. capitata and ssp. fimbriata. The interspecific embryos did not survive in vivo but could be rescued in vitro using a culture medium developed by Monnier (1973). The embryos did not grow directly into normal plants but were successfully regenerated using the protocol developed by Keller (1984). Hybridization efficiency ranged from 0 to 2.64 hybrids per pollination. Interspecific embryo abortion may be related to abnormal endosperm development.     

Comparison of callus culture with embryo culture at different times of embryo rescue for primary triticale production

Summary Callus culture of immature wheat-rye hybrid embryos was compared with embryo culture in two experiments. Embryos were rescued from field grown mother plants at two day intervals 13–21 days after pollination and plated for 1) callus culture on Murashige and Skoog medium (MS) supplemented with 2 mg/12,4-dichlorophenoxy acetic acid followed by plant regeneration on hormone free MS medium with half strength mineral salts, 2) embryo culture on Taira and Larter medium (TL). Observations were made on embryo size and condition at time of rescue (Experiment 1) and embryo development directly into plants (embryo culture) or through embryogenesis (callus culture). Fewer 19 and 21 day old embryos developed into plants from callus culture than from embryo culture in Experiment 1. Callus culture was more efficient than embryo culture in promoting plant recovery from 17 day old embryos in Experiment 2. The number of plants per embryo was significantly higher from callus culture than from embryo culture. In both experiments callus culture promoted embryogenesis in more embryos than developed in embryo culture. Embryo rescue 15–17 days after pollination was optimal in both experiments.     

红肉火龙果果皮色素的纯化及稳定性的研究

为了研究大孔吸附树脂纯化火龙果果皮色素的条件和纯化后色素的稳定性,采用静态试验和动态试验确定纯化条件,计算纯化色素的色价,并观察纯化色素在不同条件下的稳定性。结果表明：在供试的5种大孔吸附树脂中,HPD100树脂的吸附和洗脱效果最好。室温下,以30%乙醇为洗脱剂,吸附流速和洗脱流速10 mL/min时,纯化的红肉火龙果果皮色素色价为72.33,是未纯化前的5.39倍。光照不利于色素的稳定,Na+、K+、Mg2+、Ca2+、EDTA以及葡萄糖对色素的稳定性影响不大,而Sn2+、柠檬酸和酒石酸加快色素的降解。由此得出,大孔吸附树脂能有效分离纯化火龙果果皮色素,色素应避光保存,避免接触Sn2+、柠檬酸和酒石酸。     

Introgression of resistance to late blight (Phytophthora infestans) from Solanum pinnatisectum into S. tuberosum using embryo rescue and double pollination

In order to investigate the agricultural potential of the genus Vicia, and identify traits associated with productivity and responsiveness to environment, 34 undomesticated Mediterranean accessions representing Section Narbonensis (V. johannis, V. narbonensis) and V. sativa were grown in five contrasting environments in northern Syria (growing season rainfall: 76–290 mm).Highly significant genotype × environment interactions were observed for all traits. For most of the components of yield, accession mean performance (productivity)was highly correlated with responsiveness across environments (r = 0.59–0.96), as defined by joint linear regressions. Thus high yielding genotypes tended to be relatively more productive than low yielding genotypes under conditions that favoured high yields. Regression analysis revealed that mean site yields were positively correlated to rainfall (r = 0.85) and its attendant effect on growing season length as measured by cumulative season temperature and phenology (r = 0.59–0.81).In order to examine yield related traits independently of taxonomy, genotypes were grouped into three categories using K-means clustering based on productivity and responsiveness of seed, hay and biological yield. Highly productive/responsive genotypes were tall with high harvest index, large seeds and low fecundity (seeds and pods per plant), whereas unproductive/unresponsive plants tended to be short, highly fecund, with small seeds and low harvest index. Principal components analysis showed that responsiveness, in terms of seed, hay and biological yields, was closely related to phenological plasticity. Thus highly productive/responsive genotypes were able to start flowering earlier than unproductive/unresponsive genotypes in early environments, but significantly later in late, higher rainfall environments. Plant growth habit was also related to yield responsiveness. In environments with little biomass production the proportion of erect plants was high in all three categories. In more favourable, high biomass environments, the proportion of erect plants in unproductive/unresponsive genotypes fell dramatically, but was unchanged among productive/responsive genotypes. We suggest that for unproductive/unresponsive genotypes competition for light is increased under optimal growth conditions. We argue that the optimal combination of fixed and responsive traits in high yielding genotypes results in a `compound interest-type' response to more favourable environments. Highly productive and responsive genotypes can capture resources more effectively than their low yielding counterparts, leading to a positive relationship between performance and responsiveness for most components of yield. Differences in productivity and responsiveness for seed, hay and biological yield reflected Vicia taxonomy, increasing in the following order from low to high: V. johannis, V. sativa, the small seeded V. narbonensis (salmonea, jordanica, affinis) V. n. var. narbonensis, and finally V. n. var.aegyptiaca. V. n. var. aegyptiaca showed the most agricultural potential, since the taxon contained all the properties of productive/responsive genotypes listed above, yielding >1 t/ha under extremely arid conditions (104 mm),and >2.5 t/ha on 290 mm rainfall, confirming its potential for dry environments. This revised version was published online in August 2006 with corrections to the Cover Date.     

Development of triploid daylily (<Emphasis Type="Italic">Hemerocallis</Emphasis>) germplasm by embryo rescue

Interploidy crosses between diploid daylily (Hemerocallis) cultivars (2n = 22) and tetraploid cultivars (2n = 44) were performed via in vivo hybridization aiming to produce triploid hybrid germplasm (3n = 33). Plant growth regulator-free MS based medium containing 5 levels of sucrose—1, 2, 3, 4 or 5% was used to optimize the embryo rescue medium. It was determined that the medium supplemented with 3% sucrose gave highest rate of immature hybrid embryo rescue. Thirty-seven hypothetically triploid genotypes were subjected to verification of ploidy status by root tip chromosome counting and flow cytometry. Thirty-one of them were confirmed triploids. These genotypes were in vitro propagated, acclimatized to ex vitro conditions, and planted for future field performance evaluation and environmental testing. The newly developed triploid genotypes could open new horizons for further polyploidy breeding.     

Embryo rescue,for the production of F1 hybrids,in English rose

Summary The ability to recover embryos from achenes resulting from two sexual crosses of English rose varieties was investigated. Functional germination of recovered (rescued) embryos was considerably higher than for seed material when germinated conventionally. This technique also enabled the germination process to be fully controlled, and the aseptic techniques employed, avoided death of the germinating embryo due to fungal or bacterial infection. Seedlings, produced from recovered embryos, were successfully transferred to glasshouse conditions and matured to flowering plants. Many such plants exhibited combined floral characteristics of the two parent lines. The effects of media type, carbohydrate source and specifically the growth regulator BAP on the germination of recovered embryos was also assessed. The low percentage germination observed for achenes as used in this study, was due to both physical and chemical restrictions on the embryo. It is now therefore possible, to use embryo rescue techniques on English roses to recover flowering plants, of potential commercial value, that would normally be lost after sexual crossing.Abbreviations BAP 6-benzylaminopurine     

红肉火龙果(Hylocereus polyrhizus)果实与茎转录组测序分析

为探讨红肉火龙果果实和茎转录组模式及可溶性糖和甜菜苷色素积累的分子基础,采用Illumina HiSeq 4000技术对果实果肉和茎进行转录组测序。共获得28.81 Gb的Clean read,组装获得79 658个Unigene(54.98 Mb),平均长度为690 bp。35 139个Unigene能够在NR、NT、Swiss-Prot、KEGG、COG、PFAM和GO数据库中注释。大量Unigene参与多条代谢途径,包括碳水化合物和氨基酸代谢、次生代谢、柠檬酸循环、果糖和甘露糖代谢等。在果实果肉中上调表达的Unigene主要参与淀粉、蔗糖代谢途径和谷胱甘肽代谢,在茎中上调表达的Unigene主要参与碳代谢和光合作用代谢。筛选获得5个与果实可溶性糖和甜菜苷色素积累的重要候选Unigene。本研究结果克隆的火龙果果实可溶性糖和甜菜苷色素积累相关功能基因,对探讨基因的生理功能和调控机制,改良火龙果果实品质提供重要基础。     

胚胎挽救技术在油菜远缘杂交育种中的应用研究进展

本文论述了子房培养、胚珠培养和胚培养在油菜远缘杂交育种中的意义以及近年来所取得的成就,详细阐述了不同种(属)间杂交组合、培养基种类、激素种类和浓度、天然复合物、子房、胚珠和胚离体时间等影响子房培养、胚珠培养和胚培养的主要因素,并指出了胚胎挽救技术存在的主要问题和今后的研究方向.