Progeny tests of barley,wheat, and potato regenerated from cell cultures after in vitro selection for disease resistance

Summary Because plant cells cultured in vitro express genetic variability and since they can be regenerated into functional plants, procedures have been designed to use this system for the production of plants with new important agronomic characteristics, particularly for disease resistance. For barley, wheat, and potato somaclones have been found that were less susceptible to a toxin of Helminthosporium, fusaric acid, Fusarium coeruleum, F. sulphureum, or Phytophthora infestans, when screened in the first in-vitro-derived generation. Here the progeny of such somaclones is evaluated after natural and artificial infection, using greenhouse-grown or field material. The progenies of the same somaclones did not express detectable differences, which indicated that no heterozygous mutations occurred. Most lines and clones differed in their level of susceptibility to the pathogen compared to the level of the starting material, but these data were in no instance significant. It is discussed here whether this lack of significance is due to a lack of genetic differences or whether the test procedures are in adequate for detecting and securing the slight, probably quantitative, alterations.     

In vitro culture selection increases glyphosate tolerance in barley

In vitro culture of barley calluses has been used to produce plants with increased glyphosate tolerance. Calluses from immature embryos of barleyHordeum vulgare L. (Jeff) were cultured on Murashige and Skoog medium with 10^-6, 10^-5, 10^-4, 5×10^-4, 10^-3, or 10^-2M glyphosate for one, four or thirty months. Plants were regenerated from calluses maintained in glyphosate medium at 10^-6, 10^-5 or 10^-4M for four months, at 10^-5 or 5×10^-4M for one month and at 10^-5M for thirty months. The progeny of each regenerated plant was analyzed for response to glyphosate. Some progenies showed increased tolerance to glyphosate.     

Grain yield, symbiotic N2 fixation and interspecific competition for inorganic N in pea-barley intercrops

The effect of mixed intercropping of field pea (Pisum sativum L.) and spring barley (Hordeum vulgare L.), compared to monocrop cultivation, on the yield and crop-N dynamics was studied in a 4-yr field experiment using ¹⁵N-isotope dilution technique. Crops were grown with or without the supply of 5 g ¹⁵N-labeled N m^-2. The effect of intercropping on the dry matter and N yields, competition for inorganic N among the intercrop components, symbiotic fixation in pea and N transfer from pea to barley were determined. As an average of four years the grain yields were similar in monocropped pea, monocropped and fertilized barley and the intercrop without N fertilizer supply. Nitrogen fertilization did not influence the intercrop yield, but decreased the proportion of pea in the yield. Relative yield totals (RYT) showed that the environmental sources for plant growth were used from 12 to 31% more efficiently by the intercrop than by the monocrops, and N fertilization decreased RYT-values. Intercrop yields were less stable than monocrop barley yields, but more stable than the yield of monocropped pea. Barley competed strongly for soil and fertilizer N in the intercrop, and was up to 30 times more competitive than pea for inorganic N. Consequently, barley obtained a more than proportionate share of the inorganic N in the intercrop. At maturity the total recovery of fertilizer N was not significantly different between crops, averaging 65% of the supplied N. The fertilizer N recovered in pea constituted only 9% of total fertilizer-N recovery in the intercrop. The amount of symbiotic N₂ fixation in the intercrop was less than expected from its composition and the fixation in monocrop. This indicates that the competition from barley had a negative effect on the fixation, perhaps via shading. At maturity, the average amount of N₂ fixation was 17.7 g N m^-2 in the monocrop and 5.1 g N m^-2 in the intercropped pea. A higher proportion of total N in pea was derived from N₂ fixation in the intercrop than in the monocrop, on average 82% and 62%, respectively. The ¹⁵N enrichment of intercropped barley tended to be slightly lower than of monocropped barley, although not significantly. Consequently, there was no evidence for pea N being transferred to barley. The intercropping advantage in the pea-barley intercrop is mainly due to the complimentary use of soil inorganic and atmospheric N sources by the intercrop components, resulting in reduced competition for inorganic N, rather than a facilitative effect, in which symbiotically fixed N₂ is made available to barley.Abbreviations MC monocrop - IC intercrop - PMC pea monocrop - BMC barley monocrop - PIC pea in intercrop - BIC barley in intercrop     

Genetics of seedling and adult plant resistance to net blotch (Pyrenophora teres f. teres) and spot blotch (Cochliobolus sativus) in barley

Net blotch (caused by Pyrenophora teres f. teres) and spot blotch (Cochliobolus sativus) are important foliar diseases of barley in the midwestern region of the USA. To determine the number and chromosomal location of Mendelian and quantitative trait loci (QTL) controlling resistance to these diseases, a doubled haploid population (Steptoe/Morex) was evaluated to the pathogens at the seedling stage in the greenhouse and at the adult plant stage in the field. Alleles at two or three unlinked loci were found to confer resistance to the net blotch pathogen at the seedling stage depending on how progeny exhibiting an intermediate infection response were classified. This result was corroborated in the quantitative analysis of the raw infection response data as 2 major QTL were identified on chromosomes 4 and 6M. A third QTL was also identified on chromosome 6P. Seven QTL were identified for net blotch resistance at the adult plant stage and mapped to chromosomes 1P, 2P, 3P, 3M, 4, 6P, and 7P. The 7 QTL collectively accounted for 67.6% of the phenotypic variance under a multiple QTL model. Resistance to the spot blotch pathogen was conferred by a single gene at the seedling stage. This gene was mapped to the distal region of chromosome 1P on the basis of both qualitative and quantitative data analyses. Two QTL were identified for spot blotch resistance at the adult plant stage: the largest QTL effect mapped to chromosome 5P and the other mapped to chromosome 1P near the seedling resistance locus. Together, the 2 QTL explained 70.1% of the phenotypic variance under a multiple QTL model. On the basis of the chromosomal locations of resistance alleles detected in this study, it should be feasible to combine high levels of resistance to both P. teres f. teres and C. sativus in barley cultivars.     

Use of the additive main effects and multiplicative interaction model in QTL mapping for adaptation in barley

The additive main effects and multiplicative interaction (AMMI) model has emerged as a powerful analytical tool for genotype x environment studies. The objective of the present study was to assess its value in quantitative trait locus (QTL) mapping. This was done through the analysis of a large two-way table of genotype-by-environment data of barley (Hordeum vulgare L.) grain yields, where the genotypes constituted a genetic population suitable for mapping studies. Grain yield data of 150 doubled haploid lines derived from the Steptoe x Morex cross, and the two parental lines, were taken by the North American Barley Genome Mapping Project (NABGMP) at 16 environments throughout the barley production areas of the USA and Canada. Four regions of the genome were responsible for most of the differential genotypic expression across environments. They accounted for approximately 50% of the genotypic main effect and 30% of the genotype x environment interaction (GE) sums of squares. The magnitude and sign of AMMI scores for genotypes and sites facilitate inferences about specific interactions. The parallel use of classification (cluster analysis of environments) and ordination (principal component analysis of GE matrix) techniques allowed most of the variation present in the genotype x environment matrix to be summarized in just a few dimensions, specifically four QTLs showing differential adaptation to four clusters of environments. Thus, AMMI genotypic scores, when the genotypes constituted a population suitable for QTL mapping, could provide an adequate way of resolving the magnitude and nature of QTL x environment interactions.Ignacio Romagosa was on sabbatical leave from the University of Lleida and the Institut de Recerca i Tecnologia Agroalimentàries, Lleida, Spain, when this study was conducted     

A consensus linkage map of barley

A consensus linkage map of the barley genome was constructed. The map is based on six doubled haploid and one F₂ population. The mapping data for three of the doubled haploid populations was obtained via the GrainGenes database. To allow merger of the maps, only RFLP markers that produce a single scorable band were included. Although this reduced the available markers by about half, the resultant map contains a total of 587 markers including 87 of known function. As expected, gene order was highly conserved between maps and all but two discrepancies were found in closely linked markers and are likely to result from the small population sizes used for some maps. The consensus map allows the rapid localisation of markers between published maps and should facilitate the selection of markers for high-density mapping in defined regions.     

Extraction and genetic control of two new water-soluble proteins of mature barley seed

This paper describes the genetic control of two new water-soluble proteins in barley. Water-soluble proteins (WSPs) of mature barley seed form part of the albumin/globulin class of seed proteins. They can be extracted from hand-milled grain with water, though some WSPs are more efficiently extracted with a solution of 10 mM dithiothreitol. Polymorphisms for WSPs were detected in isoelectric focusing gels incorporating various ampholine combinations. Two new controlling genes (Wsp4 andWsp5) have been identified and located using wheat/barley chromosome addition lines and barley doubled haploids.Wsp4 is located on chromosome 2 (2H), andWsp5 was found to be tightly linked toWsp2 on the long arm of chromosome 7 (5HL). Segregation of a sixth gene (Wsp6) is also described, but this has not been mapped. The results are discussed with respect to other previously mappedWsp loci.This work was funed by the Scottish Office of Agriculture and Fisheries Department and the Agricultural and Food Research Council.     

Fertile transgenic barley by particle bombardment of immature embryos

Transgenic, fertile barley (Hordeum vulgare L.) from the Finnish elite cultivar Kymppi was obtained by particle bombardment of immature embryos. Immature embryos were bombarded to the embryonic axis side and grown to plants without selection. Neomycin phosphotransferase II (NPTII) activity was screened in small plantlets. One out of a total of 227 plants expressed the transferred nptII gene. This plant has until now produced 98 fertile spikes (T₀), and four of the 90 T₀ spikes analyzed to date contained the nptII gene. These shoots were further analyzed and they expressed the transferred gene. From green grains, embryos were isolated and grown to plantlets (T₁). The four transgenic shoots of Toivo (the T₀ plant) produced 25 plantlets as T₁ progeny. Altogether fifteen of these T₁ plants carried the transferred nptII gene as detected with the PCR technique, fourteen of which expressed the nptII gene. The integration and inheritance of the transferred nptII gene was confirmed by Southern blot hybridization. Although present as several copies, the transferred gene was inherited as a single Mendelian locus into the T₂ progeny.     

Changes in in vivo fluorescence quenching in rye and barley as a function of reduced PSII light harvesting antenna size

The relationship between the size of the light harvesting antenna to photosystem II (LHCII) and quenching of non-photochemical and dark level fluorescence was studied in wild-type rye (Secale cereale L. cv. Musketeer) and barley (Hordeum vulgare L. cv. Gunilla) as well as in the barley chlorophyll b-less chlorina F2 mutant (H. vulgare L. cv. Dornaria, chlorina-F2). Exposure for 10 min to an irradiance of 500 μmol m^?2 s^?1 resulted in a strong (0.71–0.73) non-photochemical (q_s) quenching of the fluorescence yield in wild-type (WT) material, while the barley chlorina F2-mutant was quenched to 75% of this level. Relaxation of q_s in darkness revealed a fast initial decay, related to relaxation of the high-energy-state dependent (q_E) part of q_s. Etiolated seedlings of rye and barley exposed to intermittent light (IML) for 36 cycles of 2 min light and 118 min darkness had suppressed Chl b and LHCII-production in both WT rye and barley, while the barley chlorina F2-mutant became totally devoid of all LHCII-polypeptides. It was found that the levels of q_s and q_s were similar in control grown barley chlorina F2 and IML-grown WT rye and barley, but q_s was reduced by 30 to 35% and q_s by 50 to 65%, respectively, as compared to control-grown. WT plants. No significant q_s could be detected in IML-grown barley chlorina F2. It is clear, from these changes in in vivo fluorescence quenching in rye and barley that a significant level of q_s is detectable even in the absence of LHCII. Only when the proximal antennae are totally absent, does q_E completely disappear. We conclude that the presence of LHCII is not an absolute requirement for q_E-quenching and suggest that distal as well as proximal antenna may contribute to q_E in vivo.