Induction of mutations for heat tolerance in potato by using in vitro culture and radiation

Heat tolerant mutants were obtained in two commercial potato cultivars, `Kufri Jyoti' and `Kufri Chandramukhi' through in vitro mutagenesis of in vitro propagated plantlets. Gamma-irradiated (20 and 40 Gy) shoots were micropropagated for three cycles (M₁V₃). A large number of the micropropagated shoots produced microtubers at 28 ^°C. Microtubers induced at high temperature had distorted shape but showed normal germination in field. Under stress conditions of high temperature, the frequency of chlorophyll variants increased in the gamma irradiation-derived material, however, nearly 40% of the plants had normal leaf tissue, whereas control plants showed completely damaged leaves. This revised version was published online in July 2006 with corrections to the Cover Date.     

Efficient hybridization between Lycopersicon esculentum and L. peruvianum via 'embryo rescue' and in vitro propagation

For the transfer of valuable traits from wild species into the cultivated tomato, excised globular-stage embryos 13 and 15 days after pollination (DAP) were cultured in vitro. Plants were regenerated from interspecific crosses of Lycopersicon esculentum cv. ‘Early pink’×L. peruvianum PI270435, and backcrosses of L. esculentum‘Giban (JF) No. 1’× (‘Early pink’× PI270435). Somatic embryos and single cotyledons emerged on hypocotyl sections of the embryos. Five to nine plantlets per embryo were obtained by clonal propagation. The hybrid nature of the plants is confirmed by comparing hybrids and parents in their ability to regenerate shoots from leaf segments in vitro, by comparing plant morphology and characteristics and by chromosome number. This study describes an efficient ‘embryo rescue’ method, as well as somatic embryogenesis by clonal propagation. A novel and simple method for the characterization of the interspecific hybrids is also reported.     

Gamma ray-induced genetic manipulations in flower colour and shape in Dendranthema grandiflorum and their management through tissue culture

Rooted cuttings of Dendranthema grandiflorum cv. ‘Puja’ were treated with different doses of gamma rays. Sectorial somatic mutations both in flower colour and shape were detected in all the doses. The original floret colour of ‘Puja’ is red‐purple and florets are flat spoon shaped. One of the mutant floret colour was yellow‐orange with original flat florets and another mutant floret colour was yellow‐orange with tubular florets. Original and mutated ray florets were cultured on agar‐solidified Murashige and Skoog basal medium supplemented with sucrose and different combinations of 1‐naphthaleneacetic acid (NAA) and 6‐benzylaminopurine (BAP). Direct shoot organogenesis was seen within 2 weeks of culture initiation. The best regeneration was obtained on medium supplemented with 1 mg/l BAP + 0.5 mg/l NAA. Shoots regenerated from all explant types were rooted in vitro and transferred to the field. Regenerated plants flowered true‐to‐explant floret colour and shape. The isolated yellow floret colour mutants and yellow floret colour mutants with tubular florets were maintained vegetatively and have proved to be true to type in two successive generations.     

Assessment of genetic and phenotypic variation among intraspecific somatic hybrids of potato, Solanum tuberosum L.

Intraspecific somatic hybrids have been produced by protoplast fusion in eight combinations involving 10 dihaploids (2n= 2x= 24) in an attempt to provide new material for potato breeding. Cytological analysis revealed extensive variation in chromosome number, such as aneuploid, aneusomatic and mixoploid hybrids. Most of the hybrids represented the expected chromosome number of 48; however, the frequency of aneuploids reached 50% in some combinations. Some hybrids carried structurally rearranged chromosomes and exhibited a high frequency of aberrant anaphases. Isozyme and random amplified polymorphic DNA (RAPD) patterns of the hybrids from the same fusion combination were uniform. In the field, somatic hybrids showed wide phenotypic variation in 20 morphological characters. There was a significant correlation between certain leaf characters and the ploidy level, which may be used to distinguish the tetraploid hybrids from hexaploids and octoploids. Tuber yield and flowering intensity were highest in tetraploid hybrids (2n= 4x= 48). Eighteen of the 73 hybrids reached higher yields than the standard variety ‘Adretta’. Floral development and fertility were restored in hybrids derived from fusions between non-flowering or sterile dihaploids.     

Callus Production and Plant Regeneration from Mature Embryos of Poa pratensis L.

Plant regeneration from callus cultures may provide a source ot somaclonal variation for the improvement ot the apomictic grass Poa pratensis L. It is first necessary to be able to induce callus and regenerate plants in this species at a high frequency. Variation was observed between 50 cutivars of Poa pratensis for callus induction and plant regeneration. Using the cultivars ‘Merion’ and ‘Victa’, three basal media were tested along with various media additives. Murashige and Skoog's basal medium with 0.2 mg 1^?1 2,4-dichlorophenoxyacetic acid, 0.1 mg 1^?1 6-benzylamanopurine, 100 mg 1^?1 casein hydrolysate and 25 g 1^?1 sucrose is considered to be a good medium for callus growth and plant regeneration. Embryo-like structures were observed in the callus of some cultivars but plant regeneration appeared to be predominantly from shoot meristems on the callus surface. The majority of regenerated shoots were green, but chlorophyll deficient shoots were obtained from media containing coconut milk. Green plantlets could be transferred to soil without difficulty.     

Evidence for phenotypic assortative mating for flower colour in periwinkle

Two experiments were conducted with periwinkle, Catharanthus roseus, to determine the extent of natural outcrossing. Three white-flowered, fully self-fertile, monogenic recessive mutants, namely, dwarf, wavy leaf margin and curved leaf were used, together with their parental white-flowered variety, ‘Nirmal’, and a normal pink-flowered variety, PS-1. The extent of total outcrossing ranged from 43.4 to 79.3% among mutants. Outcrossing between white × white-flowered plants ranged from 28.3 to 65.3% and was two to four times greater than that between white × pink-flowered plants in the three mutants. The extent of out-crossing between white × pink-flowered plants was similar “02.2-15.0%” in all mutants and also similar to that in the normal white-flowered variety,‘Nirmal’(00.4%), where white × white flower out-crossing could not be estimated. There were no large differences in the number of seeds per fruit, percentage fruit set and germination percentage of seeds obtained from self, white × white and white × pink flower crosses made in the glasshouse. There were also no significant differences in the number of flowers produced by the genotypes used in the study. The observed higher frequency of white × white flower matings compared with white × pink flower matings appeared to be due to the constancy of flower colour exhibited by the butterfly pollinators Pachliopta hector and Catopsilia pyranthae during their flower visits. Observations made on the occurrence of natural self-pollination revealed that automatic self-pollination did not occur in periwinkle.     

不同氮肥处理下水稻功能叶片的多样性分析

以52份来自水稻种质资源库的水稻品种为材料,设正常施氮和不施氮两个处理,对供试品种的功能型叶片性状进行分析。结果表明,功能叶片形态特征受到品种基因型和外部施氮水平的共同影响和调控。供试材料在不同氮肥处理下,功能型叶片形态特征上存在丰富的遗传多样性。相关性分析表明,在不同处理条件下,倒一叶长与倒二叶长和倒三叶长均呈显著或极显著正相关;倒一叶宽与倒二叶宽和倒三叶宽均呈极显著正相关。通过主成分分析,在正常施氮和不施氮条件下分别获得3个主成分,累积贡献率分别达到94.5%和90.6%,通过对各品种进行综合评价,正常施氮条件下,排前5名的水稻材料分别为滇屯502、玉米稻、雅恢2115、谷梅3号和水源258;在不施氮条件下,排前5名的水稻材料分别为玉米稻、滇屯502、巴西稻、CDR22和谷梅3号。在水稻株型育种中,应注意利用具有丰富遗传多样性的种质资源,并在亲本选配时选择遗传距离较远且综合性状表现差异较大的种质材料。     

Mutation breeding of Kalanchoë

Summary Freshly detached leaves of Kalanchoë cv. Annette and Josine were irradiated with X-rays and then planted to root and to produce adventitious plantlets. The age of the leaf, the presence or absence of the petiole as well as the genotype proved to be important factors in the production of adventitious plantlets.In Kalanchoë only solid, non-chimeral mutants were observed which indicates that the apex of the adventitious plantlets may originate from a single cell. This observation agrees with similar results in Achimenes, Saintpaulia, Streptocarpus and other species, where only a very small fraction of the mutants consisted of chimeras.The mutations observed were changes in flower colour and size, time of flowering, type of inflorescence, leaf form, size and colour, plant habit and other directly visible characters.A few promising mutants, selected by commercial plant breeders, are being propagated and will be evaluated.     

Cytogenetics and morphology of the pentaploid hybrid between Brassica napus and Orychophragmus violaceus and its progeny

A pentaploid hybrid plant (2n= 50, AACCO) between Brassica napus (AACC) and Orychophragmus violaceus (OO) showed matroclinous morphology and some patroclinous characters. Cloned progenies were mixoploid, consisting of various cells with 38–53 chromosomes, half of the cells with 50 chromosomes. The 50 chromosomes were mainly paired as 25 bivalents and segregated as 25:25 or 22:28; many other segregations were observed in some cells. Progenies produced by selfing had 38–47 chromosomes. Plants with 38 chromosomes were cytologically stable; in all other plants the chromosome number of individual cells varied between the genotype-specific maximum number and 38, indicating loss of chromosomes during mitosis. The mixoploid plants with 44 chromosomes mainly produced two kinds of mixoploid progenies with 44 and 41 chromosomes, respectively. All plants with 2n= 38 had the same morphology as the parental B. napus and were normal in fertility. These chromosome pairings and segregations in the pentaploid and its progeny support the hypothesis that O. violaceus is a natural polyploid species with a basic chromosome number of 3.     

Inheritance of morphological traits of periwinkle mutants with modified contents and yields of leaf and root alkaloids

Periwinkle Catharanthus roseus is a medicinally important plant producing anticancer and antihypertension alkaloids. Two mutants, one dwarf and one semidwarf and both, with a high alkaloid content in the roots and one mutant with a wavy leaf margin and a high alkaloid content in its leaves were obtained after induced chemical mutagenesis with ethyl methane sulphonate and N-nitroso-N-ethylurea in the variety ‘Nirmal’, which is resistant to dieback disease. These mutants were evaluated in the M₃ and M₄ generations. The dwarf and semidwarf mutants differed from the parental variety for many morphological characters, while the mutant with the wavy leaf margin differed mainly in leaf size and leaf thickness. Although both dwarf and semidwarf mutants showed a significantly higher alkaloid content in their roots in both generations, only the semidwarf mutant gave a significantly higher (23%) root alkaloid yield than the parental variety. The mutant with the wavy leaf margin showed a significantly higher alkaloid content in the leaves in both the M₃ and M₄ generations and also had a significantly higher (21%) leaf alkaloid yield than the parental variety. All three mutants were found to be controlled by monogenic recessive genes; the genes for ‘dwarfness’ and ‘semidwarfness’ were allelic to each other, with the allele for semidwarfness being dominant over the allele for dwarfness. The gene for the wavy leaf margin was inherited independently of the genes for dwarfness and semidwarfness.     

X-ray induced mutations in Begonia x hiemalis

Summary A study was made of the occurrence of mutants in plants of four clones of Begonia x hiemalis grown from leaf cuttings irradiated with 0, 1500, 2000 or 2500 rad of X-rays. Irradiation reduced the number and slowed down the development of adventitious shoots, but to a different degree in the four clones used. The percentage of mutated shoots was as high as 18–35% in the non-irradiated shoots. Irradiation raised it to more than 80%. In the control plants, almost exclusively flower colour mutants were found. Irradiation caused a wider range of flower colour mutants and also caused mutations of growth habit, colour and size of the leaves, and size and shape of the flower. Almost all mutants were solid, i.e. non-chimeral.Publication 405, Department of Horticulture, Agricultural University, Wageningen, the Netherlands.     

Intergeneric hybrids between Chrysanthemum morifolium ‘Nannongxiaoli’ and Artemisia vulgaris ‘Variegata’ show enhanced resistance against both aphids and Alternaria leaf spot

Intergeneric hybridization is an effective way of improving crop resistance against biotic and tolerance to abiotic stresses. Here, we describe an intergeneric hybrid between Chrysanthemum morifolium ‘Nannongxiaoli’ and Artemisia vulgaris ‘Variegata’ using ovule rescue. Chromosome counting confirmed that out of the 124 regenerated plantlets, 19 were genuine hybrids, carrying the expected hybrid number of somatic chromosomes (45, representing 27 from ‘Nannongxiaoli’ and 18 from ‘Variegata’). The 19 genuine hybrids were vigorous, flowered normally and possessed the cut flower characteristics of ‘Nannongxiaoli’. The hybrids were more resistant against both aphid infestation and Alternaria leaf spot inoculation than the ‘Nannongxiaoli’ parent, but less than the ‘Variegata’ parent.     

Studies on resistance to bacterial speck (Pseudomonas syringae pv. tomato) in tomato cv. Ontario 7710

Plants of 17 tomato cultivars and four wild Lycopersicon accessions were evaluated for their reaction to Pseudomonas syringae pv. tomato (Pst) in a greenhouse following a leaf‐spray inoculation. The genotypes exhibited a large amount of variation in response to Pst infection, with disease severity index (DSI) ratings from 0.2 to 3.9. The cultivar ‘Ontario 7710’ and two accessions of Lycopersicon hirsutum (LA 1773 and LA 1775) were the most resistant, with DSI values of 0.2, 0.4 and 0.6, respectively. Three varieties, M 1812, Kujawski and Warszawski, also showed a high level of tolerance. The most susceptible was ‘A 100’(DSI = 3.9). The inheritance of resistance to bacterial speck was investigated by disease tests in segregated populations obtained by hybridizing the tomato cv. Ontario 7710 with the susceptible variety ‘A 100′. Plants were rated for disease severity by inspecting each plant and were then evaluated according to phenotypic similarity to ‘Ontario 7710’ or ‘A 100’ in respect of the number and size of the spots. Genetic analysis in F1, F2 and backcross segregations indicated that resistance of'Ontario 7710’ to Pst is conferred by one incompletely dominant gene, Pto.     

杂种小叶杨叶片外植体离体再生体系的建立

为了开发新的小叶杨基因保存技术并提高其抗病虫害能力,本研究以杂种小叶杨(Populus simonii Carr.× P. deltoides cv. ‘Shanhaiguanensis’)无性系的叶片作为外植体,研究了杂种无性系的的离体培养及叶片再生体系,探讨了不同激素组合对杂种无性系不定芽的诱导、分化以及生根的影响。结果表明：最适于杂种小叶杨叶片分化的培养基是MS+0.1 mg/L 6-BA+0.04 mg/L NAA+0.005 mg/L TDZ,不定芽分化率和平均分化芽芽数分别达到最高的90%和6.8,芽长为2.14 cm;最佳生根培养基为1/2 MS+0.1mg/L IBA,生根率达90%。最后将这些再生植株成功驯化并移栽到温室。10 mg/L的卡那霉素可以抑制杂种小叶杨的诱导分化,20 mg/L的卡那霉素可以抑制杂种小叶杨不定芽的生根。本研究结果将有助于小叶杨抗病虫害等方面的遗传转化及相关研究,同时也将为小叶杨基因库的保存提供重要技术支撑。     

Barley lines showing prominent high green shoot regeneration in cultures derived from immature embryos

Eighty‐four cultivars of barley (Hordeum vulgare) and 95 wild strains (82 of H. spontaneum and 13 of H. agriocrithon) were surveyed for the production of callus, callus growth, and shoot regeneration in cultures derived from immature embryos. All cultivars except for ‘Turkey 381′, induced calli from more than 90% of embryos. On the other hand, the wild lines showed a large variation in the percentage of callus induction from 0 to 100%. Among the cultivars, those with the brittle rachis genotype, bt Bt₂, on chromosome 3H, regenerated shoots with a significantly higher percentage than the cultivars with the Bt bt₂ genotype. Green shoots were produced in a higher ratio (0.84) in the cultivars than in the wild lines (0.52). Among the lines examined,‘Lenins’ regenerated shoots efficiently (90.4%) and produced the highest number of calli with green shoots per embryo (4.77) followed by ‘Golden Promise’ (3.15). Examination of callus growth and shoot regeneration from embryos at different developmental stages revealed that scutellum development affected the quantity and quality of callus and shoot regeneration.     

Broad-few-leaflets and outwardly curved wings: two new mutants of chickpea

This study was aimed at the induction of morphological mutations for increasing genetic variability and making available additional genetic markers for linkage studies in chickpea (Cicer arietinum L.). A wilt‐resistant, well‐adapted chickpea cultivar of central India,‘JG 315’(Jawahar gram 315), was used for the induction of mutations. Seeds presoaked in distilled water for 2 h were treated with ethyl methane sulphonate (EMS) using six different concentrations (0.1, 0.2, 0.3, 0.4, 0.5 and 0.6%) and two different durations (6 and 8 h). Several morphological mutants were identified in M₂. One of the mutants, isolated from a treatment of 0.3% EMS for 8 h, had five to nine large leaflets per leaf in comparison with 11‐17 normal‐sized leaflets per leaf observed in the parental cultivar ‘JG 315′. The mutant was designated broad‐few‐leaflets. Many leaves of this mutant showed a cluster of three to five overlapping leaflets at the terminal end. The other mutant, designated outwardly curved wings, was isolated from the 0.5% EMS treatment for 6 h. In this mutant, the wings were curved outwards, exposing the keel petal, while the wings in typical chickpea flowers are incurved and enclose the keel. The lines developed from the broad‐few‐leaflets and outwardly curved wings mutants were named JGM 4 (Jawahar gram mutant 4) and JGM 5, respectively. Inheritance studies indicated that each of these mutant traits is governed by a single recessive gene. The gene for broad‐few‐leaflets was designated bfl and the gene for outwardly curved wings was designated ocw. The locus bfl was found to be linked with the locus lg (light green foliage) with a map distance of 18.7 ± 6.3 cM.     

Identification and mapping of adult‐plant stripe rust resistance in soft red winter wheat cultivar ‘USG 3555’

Little is known about the extent or diversity of resistance in soft red winter wheat (Triticum aestivum L.) to stripe rust, caused by the fungal pathogen Puccinia striiformis f.sp. tritici. The soft red winter (SRW) wheat cultivar ‘USG 3555’ has effective adult‐plant resistance to stripe rust, which was characterized in a population derived from ‘USG 3555’/‘Neuse’. The mapping population consisted of 99 recombinant inbred lines, which were evaluated for stripe rust infection type (IT) and severity to race PST‐100 in field trials in North Carolina in 2010 and 2011. Genome‐wide molecular‐marker screenings with 119 simple sequence repeats and 560 Diversity Arrays Technology (DArT) markers were employed to identify quantitative trait loci (QTL) for stripe rust resistance. QTL on chromosomes 1AS, 4BL and 7D of ‘USG 3555’ explained 12.8, 73.0 and 13.6% of the variation in stripe rust IT, and 13.5, 72.3 and 10.5% of the variation in stripe rust severity, respectively. Use of these and additional diagnostic markers for these QTL will facilitate the introgression of this source of stripe rust resistance into SRW wheat lines via marker‐assisted selection.     

Development of early-flowering Kabuli chickpea with compound and simple leaves

Terminal drought is a major constraint to chickpea (Cicer arietinum L.) production. Autumn sowing and early flowering have been suggested as ways to benefit from the winter rains in short rainy seasons under dryland cropping. High‐yielding, late‐flowering, simple‐leafed (slv/slv) chickpea cultivars with good field resistance to Ascochyta blight have been bred recently. Changing plant architecture, by altering leaf shape, may affect agronomic performance. As no information is available on the effect of leaf shape on phenology and seed yield, this study was aimed at: (i) introducing the simple leaf trait into an early‐flowering chickpea background; (ii) comparing the grain yield of the two leaf types in early vs. late flowering backgrounds and (iii) producing breeding lines combining early flowering, large seeds and Ascochyta tolerance with both leaf types. Hybrid progeny were studied from the cross of ‘Sanford’ (slv/slv) and ICC7344, (compound, SLV/SLV). Four early‐podding, F₈ breeding lines were selected with either simple or compound leaves. In three different field experiments under dryland conditions (334–379 mm), they yielded ca. 1.4 t/ha as compared with 1.0 t/ha in the standard Israeli ‘Yarden’ on one site, but no significant differences in yield were obtained in the other two experiments.     

Callusing and Regeneration of Plantlets via Somatic Embryogenesis from Inflorescence Cultures of Triticum aestivum L.: Role of Genotype and Long-Term Retention of Morphogenic Potential

In the present investigation we have succeeded in obtaining a high frequency of regeneration of plantlets via somatic embryogenesis from callus derived from immature inflorescence explains of Triticum aestivum var, ‘Sonalika’. The explants were cultured on MS medium supplemented with 2,4-D, casein hydrolysate and coconut milk. A large number of embryoids germinated, to form plantlets on the medium when 2, 4-D was omitted altogether or provided at low concentration. Plantlets were transferred to soil under natural environmental conditions and were shown to have the normal chromosome number of 2n = 6x = 42. Experiments with nineteen other varieties show that there is a marked effect of genotype both on initiation of callusing as well as on regeneration. So far ‘Sonahka’ has proved to be the most responsive among varieties tested by us. With callus of the variety ‘Sonalika.’, we also conducted an investigation on long-term retention of regenerative potential. During Song-term culture, for about 12 months, the morphogenic potential gradually diminished and was finally lost, but the regeneration potential could be restored by subculturing at very short intervals.     

绿豆EMS诱变突变体库的构建及表型分析

为探索适于绿豆突变体库构建的最佳甲基磺酸乙酯(EMS)浓度和处理时间,以‘皖科绿3号’种子为研究材料,设置4种EMS浓度（0.6%,1.0%,1.4%和1.8%）和4种时间（6 h,10 h,14 h和18h）共16个组合,并以半致死剂量为突变体库构建的条件。结果表明,‘皖科绿3号’的最佳诱变条件为1.0%/10 h。用该条件处理6000粒成熟的‘皖科绿3号’种子,获得M₁代植株998份。对M₂代群体中植株叶色、叶形、株高、花器官、开花期等性状进行鉴定。共发现表型变异材料324株,变异频率为2.97%。突变体中叶色变异和矮秆是主要的变异类型,分别占植株总数的1.10%和0.46%。利用基因组重测序技术对4份突变材料的突变位点进行鉴定,测序结果表明碱基突变频率为1/37.78 kb。实验确定‘皖科绿3号’的最佳诱变条件为1.0%/10 h。研究结果也表明利用EMS诱变的方法可获得丰富的表型变异材料,可用于绿豆品种遗传改良和功能基因组学研究。