High-performance liquid chromatographic analysis of anthocyanins in Japanese garden iris and its wild forms

Summary Following bud pollination of tartary buckwheat (Fagopyrum tataricum) with pollens from common buckwheat (F. esculentum), cv. Mancan, 31% of the ovaries began to grow but all turned brown and withered after 6 to 14 days. Fluorescence microscopy of the growing ovaries showed the pollen-tube entering the embryo-sac. The ovules from the growing ovaries failed to produce any embryo in the culture medium in which the immature embryos from the self-pollinated (compatible) tartary flowers were able to mature and germinate. No embryo development was observed after cross-pollination. The interspecific incompatibility is attributed to the failure of gametes to fuse.Abbreviations BAP benzylaminopurine - IAA indoleacetic acid     

Diploid and tetraploid progenitors of wheat are valuable sources of resistance to the root lesion nematode <Emphasis Type="Italic">Pratylenchus</Emphasis><Emphasis Type="Italic">thornei</Emphasis>

The root lesion nematode Pratylenchus thornei is widely distributed in Australian wheat (Triticum aestivum) producing regions and can reduce yield by more than 50%, costing the industry AU$50 M/year. Genetic resistance is the most effective form of management but no commercial cultivars are resistant (R) and the best parental lines are only moderately R. The wild relatives of wheat have evolved in P. thornei-infested soil for millennia and may have superior levels of resistance that can be transferred to commercial wheats. To evaluate this hypothesis, a collection of 251 accessions of wheat and related species was tested for resistance to P. thornei under controlled conditions in glasshouse pot experiments over two consecutive years. Diploid accessions were more R than tetraploid accessions which proved more R than hexaploid accessions. Of the diploid accessions, 11 (52%) Aegilops speltoides (S-[B]-genome), 10 (43%) Triticum monococcum (A ^m -genome) and 5 (24%) Triticum urartu (A ^u -genome) accessions were R. One tetraploid accession (Triticum dicoccoides) was R. This establishes for the first time that P. thornei resistance is located on the A-genome and confirms resistance on the B-genome. Since previous research has shown that the moderate levels of P. thornei resistance in hexaploid wheat are dose-dependent, additive and located on the B and D-genomes, it would seem efficient to target A-genome resistance for introduction to hexaploid lines through direct crossing, using durum wheat as a bridging species and/or through the development of amphiploids. This would allow resistances from each genome to be combined to generate a higher level of resistance than is currently available in hexaploid wheat.     

Marker assisted backcross breeding approach to improve blast resistance in Indian rice (Oryza sativa) variety ADT43

Marker assisted backcrossing breeding has become one of the essential tools in transferring novel genes to adapted varieties and was employed to pyramid three blast resistance genes Pi1, Pi2 and Pi33 to a popular susceptible rice variety ADT43. Gene pyramiding process was facilitated by marker aided selection for both foreground as well as background genotype. Previously reported linked molecular markers were deployed to survey resistant and susceptible genotypes. In the BC₃F₁ generation four lines viz, AC-B3-11-7, AC-B3-11-36, AC-B3-11-57, AC-B3-11-83 were identified to be pyramided with three genes and subjected to background analysis and a genome recovery up to 95 % was observed and advanced to further generations. Morphological, yield and grain quality traits were significantly similar to ADT43. The introgressed lines with three gene combinations were highly resistant to the blast pathogen compared to genotypes with single genes and the susceptible checks under blast nursery screening at two epiphytotic locations; Coimbatore and Gudalur. The selected three gene pyramided backcross lines in the desirable background were advanced to obtain an improved ADT 43 with resistance to blast disease.     

Effect of in vitro propagation methods on field performance of two strawberry cultivars

Summary Strawberry cultivars, Redcoat and Veestar, propagated by meristem culture (MC), callus culture (CC) and direct shoot regeneration (DR) from leaf disks were compared for their vegetative and reproductive characters with standard runner (SR) propagated plants under field conditions. In the planting year, in vitro propagated plants of both cultivars had the same number of leaves as SR plants, but in vitro propagated Redcoat produced fewer stolons per plant than SR plants. However, in the following year, in vitro propagated mother plants of both cultivars had more leaves and higher runner production than SR mother plants. Flowering and fruiting behaviour of Veestar was not appreciably influenced by in vitro propagation methods. However, in vitro propagated plants of Redcoat flowered earlier and produced more flowers and fruits than SR plants, but still maintained normal berry weight. Among in vitro propagated plants, DR plants of Redcoat were the earliest to flower, whereas MC plants produced more flowers and fruits. The field performance of the first daughter plants derived from the in vitro propagated plants was consistent with their respective mother plants. Leaf shape of both cultivars was not altered by in vitro propagation. Phenotypic abnormalities were mainly confined to occurrence of yellow leaf variants in MC and CC plants and occasional appearance of plants with irregular flowering and growth habit among CC plants.NRCC No. 38004     

Race 1 fusarium wilt tolerance on banana plants selected by fusaric acid

Summary The selection of tolerant variants to race 1 fusarium wilt of banana was carried out through the effects of fusaric acid onin vitro banana multiple bud clumps (MBCs). The MBCs of Maçã cultivar (Musa sp., AAB, Silk), which is susceptible to the race 1 fusarium wilt, were used. And, Nanicão cultivar (AAA, Cavendish subgroup) was used in the tolerant tests as control of a disease tolerant variety. Firstly, to aim at determining an appropriate concentration of the toxin for the tolerant selection, the MBCs were cultured on the Modified Murashige & Skoog medium supplemented with 0.05 to 1.6 mM fusaric acid. The growth on both cultivars was completely inhibited on the medium containing 0.1 mM fusaric acid.Tolerant variants of Maçã were, then, selected with 0.1 mM fusaric acid after chemical mutagen treatment. The results showed an increased tolerance of the selected Maçã plants to the race 1 fungus in greenhouse tests. In vitro selection by fusaric acid is a very useful method for obtaining fusarium disease tolerance, although the tolerance mechanism of the selected plants may be different from that of existing tolerant cultivars.     

High frequency somatic embryogenesis with a pampeana-derived genotype of alfalfa (Medicago sativa L.)

Summary Somatic embryos of genotype R11 of the alfalfa variety Pampeana were produced from embryogenic calli derived from leaf sections. They were induced by an auxin shock and its development was attempted on six different media. The best condition for somatic embryo production was inducing callus on MS medium plus 10 M 2,4-D and 4,6 M KIN and transferring them, after the auxin shock, to MS with 10–20 mM NH₄ ⁺ and 30 mM proline. More than 500 somatic embryos per plate were produced. Embryos were grown to plants on MS or half strength MS media and all regenerated plants resembled the original R11 genotype. This technique could be useful in alfalfa Pampeana improvement using genetic modification.     

Adaptability and variation in <Emphasis Type="Italic">Isatis tinctoria</Emphasis> L.: a new crop for Europe

Isatis tinctoria L. was cultivated until the 19th century to produce indigo, a natural blue pigment used principally for dyestuffs. The current search for alternative crops and interest in natural products has led to reconsidering I. tinctoria as a crop to be grown in marginal areas to produce natural indigo. To reintroduce I. tinctoria into cultivation, its behaviour under different climatic conditions as well as its morpho-physiological and genetic diversity must be assessed in order to evaluate the possibilities of future breeding work. To do this, a Eurasian collection of 15 accessions was studied in a 2-year experiment. The study was carried out in four locations in order to assess plant performance at altitudes ranging from 380 to 1,700 m a.s.l. A second experiment evaluated the morpho-physiological diversity of several traits (some related to agronomic performances) of the collection. In a third experiment the genetic traits of the collection were characterised by using eight AFLP and eight SAMPL markers. The species showed a wide adaptability to different mountainous conditions and the populations showed high morphologic and genetic variability and differed according to their origins. Both morpho-physiological and molecular characterisation allowed the accessions to be distinguished into groups of European and Asian origin. Future breeding work is recommended because some accessions have good agronomic potential.

Interspecific hybrids of Cucumis metuliferus × C. anguria obtained through embryo culture and somatic embryogenesis

Summary Interspecific crosses between Cucumis metuliferus Naud. and C. anguria L. were obtained through embryo culture. Embryos in the rabbit-ear to advanced fluke-shaped stages were rescued 34–99 days after pollination. Plants were obtained through direct embryo culture, and through somatic embryogenesis from immature embryos. For direct embryo culture, fluke-shaped embryos were stored in sterile water in darkness for three days at 25C prior to transfer on Murashige and Skoog (MS) culture medium plus 1.0 M 6-benzylamino-purine. Multiple plants were obtained from single embryos through somatic embryogenesis of rabbit-ear stages on MS plus 10 M indole-3-acetic acid and 5 M 6-benzylamino-purine. Evidence of hybridization included leaf shape intermediate between the two parents, penduncle shape prior to fertilization which resembled the male parent, low pollen viability and isoelectric focussing of protein bands for acid phosphatase of leaf extracts.     

Genotypic and exogenous factors affecting shoot regeneration from anther callus of linseed (Linum usitatissimum L.)

Summary The objective of this study was to investigate factors affecting the regeneration capacity of linseed anther culture. Four different environmental conditions in a phytotron were tested with regard to their effects on anther donor plants of cv. Hella. Anther response and shoot regeneration from anther callus was maximal when donor plants were grown in a 16 hrs-day at 14°C day/8°C night temperature. Anthers of four linseed genotypes were cultured on different media. Maximum shoot regeneration was achieved when the induced calli were transferred onto a modified N6 medium containing zeatin (1 mg l^-1). Most of the calli regenerated shoots in the second subculture on regeneration media. Shoots were rooted on modified B5 or MS media containing NAA (0.1 mg l^-1). Cytological examinations of incubated anthers and root tips of regenerated plants indicated that the anther calli were derived from microspores.Abbreviations B5 Gamborg's (1975) medium - BAP 6-benzylaminopurine - 2,4D dichlorophenoxyacetic acid - N6 Chu's (1978) medium - NAA -naphthaleneacetic acid - MS Murashige & Skoog's (1962) medium - ZEA zeatin     

Production of intergeneric hybrid plants between <Emphasis Type="Italic">Sandersonia aurantiaca</Emphasis> and <Emphasis Type="Italic">Gloriosa rothschildiana</Emphasis> via ovule culture (Colchicaceae)

Intergeneric hybrid plants between Colchicaceous ornamental plants, Sandersonia aurantiaca and Gloriosa rothschildiana, have successfully been produced via ovule culture. After 5 days of reciprocal cross-pollination, a few pollen tubes were observed in the ovary. Although seeds were obtained in both reciprocal cross-combinations, they did not germinate under ex vitro conditions. Ovules with placental tissues isolated 14 days after cross-pollination of S. aurantiaca × G. rothschildiana were cultured on a medium containing 0.01 mg l^–1 each of -naphthaleneacetic acid (NAA) and 6-benzyladenine (BA), on which 41.5% of ovules swollen and produced callus-like structures within 10 weeks. When such swollen ovules were transferred to a medium containing 0.1 mg l^–1 each of NAA and BA, 7.5% of the initially cultured ovules produced rhizome-like structures within 6 weeks. Among the rhizome-like structures, those derived from two independent ovules (3.7% of the initially cultured ovules) produced multiple shoots following transfer to a medium containing 0.25 mg l^–1 NAA and 2.5 mg l^–1 BA. Multiple shoot-derived plantlets were established on a plant growth regulator-free medium, and they were successfully transplanted to pots. Early verification of their hybridity was accomplished by flow cytometry (FCM) analysis, chromosome observation and rDNA analysis.     

Bolting resistant breeding of Chinese cabbage. 1. Flower induction of late bolting variety without chilling treatment

Summary When a local slow bolting variety Osaka Shirona Bansei (Brassica rapa L. ssp. pekinensis, syn. B. campestris L. ssp. pekinensis) was grown in a phytotron (25°C, 16 hours day length without chilling treatment), one third of the plants bolted and flowered. In order to clarify the different flowering responses in the variety, a progeny line (FNC) of the flowering plants was chilled for 4 different periods (0, 22, 36 and 53 days) in a chamber of 2 7°C, then transplanted to three different conditions, i.e. PHY: 25–20°C day and night temperatures, 16 hours day length, GHL: 10 25°C, 16 hours day length with supplementary light and GHN: 10 25°C, natural day length (10 15 hours). In PHY, FNC bolted and flowered with almost the same leaf numbers in all 4 different chilling treatments. This means that FNC has very low sensitivity and no requirement to low temperature for its reproductive growth. In GHN (short day length), FNC bolted very slowly. Then the bolting and flowering of FNC were promoted by both long day length and high temperature. The newly found bolting characteristics of Osaka Shirona Bansei could be applied to breed unique slow bolting Chinese cabbage (B. rapa L. ssp. pekinensis) which might be non-sensitive to low temperature and its bolting and flowering would be induced with the combination of long day length and high temperature. Using the unique variety, it might be also possible to establish a new cropping type of Chinese cabbage (late autumn sowing, spring harvest).Abbreviations PHY Phytotron, long day and high temperature condition - GHL Greenhouse, long day and medium temperature condition - GHN Greenhouse, natural (short) day and medium temperature condition - FNC A progeny line of Osaka Shirona Bansei which flowered with no chilling treatment - FC A progeny line of Osaka Shirona Bansei which flowered with chilling treatment     

In vitro production of male sterile Zinnia elegans

Summary Axillary bud proliferation of Zinnia elegans Jacq. was accomplished from vegetative bud explants of greenhouse-grown plants. Photoperiod extension via metalarc lamps in the greenhouse stock plant environment significantly increased both the quantity and quality of explants. Shoot cultures initiated from axillary buds of identified male sterile plants proliferated best on a medium with 1M BAP. Microshoots exhibited 80% rooting on media without growth regulators, and 100% acclimatization of rooted plantlets was accomplished in a lighted, high humidity chamber. In vitro- produced male sterile plants were more compact, more heavily branched, and demonstrated earlier flowering than seed-propagated plants. These male steriles were cross pollinated to male fertile lines to verify their ability to serve as a uniform female parent.     

Associations between three ml-o powdery mildew resistance genes and agronomic traits in barley

Summary A population of 198 chromosome-doubled haploid lines of spring barley was scored for segregation in locus ml-o (powdery mildew reaction) on chromosome 4 and in the linked loci s (rachilla hair length) and ddt (reaction to the insecticide DDT) on chromosome 7. They were also tested in a disease-free field trial for the agronomic traits: grain yield, thousand grain weight, lodging, and necrotic leaf spotting. The three mutagen-induced resistance genes ml-o5, ml-o6 (from Carlsberg II) and ml-10 (from Foma) showed no detectable differences with respect to effects on agronomic traits. They all conferred a four per cent reduction in grain yield caused mainly by lower thousand grain weight, and an increase in necrotic leaf spotting. The two original mutants of Carlsberg II had additional mutant genes affecting agronomic traits. Lines with gene S (long hair) had on average a three per cent higher thousand grain weight than those with s. The alleles in locus ddt showed no association with the agronomic traits. It is concluded i) that the associations between the three ml-o alleles and agronomic traits are caused by pleiotropy, ii) that ml-o resistant, high-yielding lines may be selected, and iii) that the association between gene s and thousand grain weight may be due to genetic linkage.Abbreviations DH-lines chromosome-doubled haploid lines     

Characterization of the resistance to <Emphasis Type="Italic">Phytophthora infestans</Emphasis> in local potato cultivars in Bolivia

This experiment was carried out to investigate whether and how much field resistance to late blight, caused by Phytophthora infestans, is present in the local cultivated potato germplasm. In total 36 entries were compared in a field experiment in an area highly conducive to late blight development. Of the 36 cultivars 32 were local cultivars belonging to five Solanum species, S. tuberosum (1 accession), S. andigena (18), S. juzepczukii (2), S. stenotomum (9) and S. ajanhuiri (2). The other four cultivars were derived from breeding programmes, one being the Dutch cultivar Alpha used as a highly susceptible control. The 36 cultivars were planted according to a simple 6 × 6 lattice design with three replicates. Each replicate was divided in six incomplete blocks each with six cultivars. The disease severity was assessed weekly during 9 weeks starting 48 days after planting. The area under the disease progress curve (AUDPC) was used as a measure of the field resistance. Nine isolates from surrounding potato fields were tested for their virulence to the resistance genes R1–R11 using 22 differential cultivars. The components of the field resistance of 19 of these cultivars were compared in the greenhouse using a local isolate with virulence to all known R-genes, except to R9. The nine isolates represented seven races with a race complexity varying from 7 to 10 virulence factors. All isolates carried virulence against R1, R2, R3, R7, R10 and R11, while virulence against R9 was absent. The AUDPC among the 32 local cultivars ranged from very large, significantly larger than that of ‘Alpha’ to very small. The AUDPC from S. stenotomum accessions ranged from very large to intermediate, those from S. andigena accessions from large to very small. Especially among the S. andigena accessions interesting levels of field resistance were found. Four components of field resistance were assessed, latency period (LP), lesion size (LS), lesion growth rate (LGR) and relative sporulation area (RSA). All four showed a considerable variation among the cultivars. The LP ranged from 3½ to 6 days. The LS ranged from 225 mm² to 20 mm². The LGR varied about six-fold, the RSA more than 10-fold. The components tended to vary in association with one another. LP and LGR were well associated with each other and had a significant correlation with the AUDPC.     

Analysis of Chinese Spring regenerants obtained from short- and long-term wheat somatic embryogenesis

Summary Somatic embryogenesis was initiated from immature embryos on Murashige-Skoog (MS) medium plus 2 mg.l^-1 2,4-dichlorophenoxyacetic acid, 2% sucrose and 0.6% agarose. Somatic embryos were isolated and regenerated into whole green plants on MS medium devoid of 2,4-D. These regenerants were previously demonstrated to differ in their mitochondrial DNA organization. In order to estimate their characteristics three progenies of short-term culture regenerants and three progenies of long-term culture regenerants were analyzed and compared to the parental line. These somaclones obtained from the wheat variety Chinese Spring were evaluated for variation of 13 agronomic and morphological quantitative characters in comparison to the parental line. Significant variation was observed for plant height, spike length, main tiller diameter, between the somaclones regenerated from long-term culture and their parent. Differences were observed to increase with the duration of culture, leading to a significant modification of the structure of the plants. Several changes occurred during the somatic tissue cultures, but to a lesser extent than has previously been described in the literature.     

Hybridization barriers between wheat and rye: In vitro pollen assays and electrophoretic survey

Summary Five- to seven-day-old ovaries from Triticum aestivum vars Hope (non-crossable), Chinese Spring (CS, crossable) and the chromosomal substitution line Chinese Spring/Hope 5B (CS/H5B) were used to test the hypothesis that an ovarian substance inhibits rye (Secale cereale var Wrens) pollen tube growth. In vitro assays on a semi-solid medium demonstrated that a soluble, dialyzed lysate from Hope or CS/H5B ovaries inhibited rye pollen tube elongation significantly more than a similar lysate from CS ovaries (26%, 13% and 5% respectively compared to controls). When only the 20 longest tubes from each sample were included in the analysis the relationship was maintained (33%, 22% and 15% for Hope, CS/H5B and CS respectively). Heating the Hope lysate at 100° C for 10 min reduced the inhibition from 26% to 14% for all pollen tubes measured (n = 51 per sample) and from 33% to 20% when only the 20 longest pollen tubes per sample were analyzed. Isoelectric focusing followed by sodium dodecyl sulfate polyacrylamide electrophoresis (IEF/SDS-PAGE) and non-equilibrium pH gradient electrophoresis (NEPHGE) revealed several differences in the polypeptide profiles of soluble lysates from vars Hope, CS and CS/H5B. While two striking differences were observed, a 50 kilodalton (kD) polypeptide with an isoelectric point (pI) of approximately 8.5 and 100 kD polypeptide (pI4) from CS lysates which were considerably reduced in lysates of Hope and CS/H5B, none were obviously associated with the Hope 5B chromosome and therefore cross-incompatibility.Mention of a trademark or proprietary product is for identification only and does not imply a guarantee or warranty of the product by the U.S. Department of Agriculture.     

Heritable tissue culture induced variation in Zinnia marylandica

Summary Adventitious shoots of Zinnia marylandica, an amphidiploid with limited genetic segregation, were regenerated from cotyledonary tissue on Murashige-Skoog (MS) media containing 0.2 or 22.2 M thidiazuron (TDZ) and grown through flowering. Fisher's Test for Equal Variance indicated tissue culture induced plants had more variation than seed-derived control plants. Twelve of 149 (8%) plants derived from 0.2 M TDZ and three of 23 (13%) plants from 22.2 M TDZ had variant characters. Aberrant characteristics in self-pollinated variants included plant height, fertility, flower color and morphology, and were sexually transmitted, indicating genetic change had occurred. Aberrant characteristics not observed in regenerated plants arose in progeny.Abbreviation TDZ thidiazuron     

The effect of colchicine on triticale anther-derived plants: Microspore pre-treatment and haploid-plant treatment using a hydroponic recovery system

In cereals, chromosome doubling of microspore-derived haploid plants is a critical step in producing doubled haploid plants. This investigation was undertaken to study the effect of incorporation of colchicine in the induction medium for anther culture, and the effect of colchicine on anther culture-derived plants of triticale grown under controlled greenhouse conditions. In the latter case, chromosome doubling of adult sterile plants derived from anther culture of fourteen triticale populations was attempted, where androgenetic plants with non-dehiscent anthers were cloned and subjected to the colchicine treatment, and then grown with the aid of hydroponics. The hydroponic system provided optimal conditions for recovery of the affected haploids from the toxic effects of colchicine treatment and all colchicine-treated plants survived. A topcross-F₁ (TC₁F₁) population with timopheevii cytoplasm produced the highest percentage of plants with seed-set either due to chromosome doubling by colchicine (98%) or spontaneous doubling of chromosome number (15%). Colchicine-treated anthers performed inferior than control in both induction and regeneration phases. One of the key observation of this study was the reversal from reproductive stage back to the vegetative stage which in turn enabled further cloning of haploid plants under hydroponic conditions once they were identified as sterile. The one hundred percent survival rate of in vitro-derived plants, 100% survival rate of colchicine treated haploid plants and the high chromosome doubling success rate (X = 82.3) observed in this study imply that a temperature-controlled greenhouse with an hydroponic system provides an efficient environment for inducing chromosome doubling of haploid plants in cereals. This revised version was published online in August 2006 with corrections to the Cover Date.     

Stability of bacterial leaf spot resistance in peach regenerants under in vitro,greenhouse and field conditions

Summary Phenotypic stability of bacterial leaf spot resistance in peach (Prunus persica (L.) Batsch) regenerants, either selected at the cellular level for insensitivity to a toxic culture filtrate of Xanthomonas campestris pv. pruni or screened at the whole plant level for resistance to X. campestris pv. pruni, was investigated. A detached-leaf bioassay was used to evaluate the original regenerants again after three years in the greenhouse and also after a two to three year cycle of tissue culture propagation. Peach trees derived through micropropagation from the original regenerants were also evaluated after one to three years growth in the field. Although leaf spot resistance was retained in some regenerants over time in the greenhouse, following in vitro propagation, and under field conditions, resistance was either lost or not expressed in others. Regenerants # 19-1 and #156-6, derived from embryo callus of bacterial spot susceptible Sunhigh, were significantly more resistant than Sunhigh. High levels of resistance were exhibited in greenhouse plants and field-grown trees of regenerant #122-1, derived from embryo callus of moderately resistant Redhaven. This research provides additional evidence that selecting or screening for somaclonal variants with disease resistance is a feasible approach to obtaining peach trees with increased levels of bacterial spot resistance.Abbreviations TC Tissue-Cultured - TF Toxic culture Filtrate     

The black currant leaf curling midge,Dasyneura tetensi Rübs.; its host range,and the inheritance of resistance

Summary Established Western European black currant cultivars are susceptible to Dasyneura tetensi but the Scandinavian Sunderbyn II, Kangosfors and Hedda, and 10 out of 33 Russian cultivars showed very slight or no leaf symptoms of midge attack in the field.Of species within the black currant Section Eucoreosma, Ribes dikuscha, R. pauciflorum (probably=syn R. ussuriense), and R. americanum were not attacked, but an accession of R. bracteosum proved very susceptible. Species within other sections of the genus were not attacked.A strong dominant resistance gene in R. dikuscha, designated Dt, appeared to be linked with (a) gene(s) controlling resistance to American gooseberry mildew, Sphaerotheca mors-uvae. Resistance in Sunderbyn II and, probably, in R. pauciflorumwas oligogenic and dominant.