Relationship between somaclonal variation and type of culture in cucumber

Highly inbred B line of cucumber was used to compare the effect of four types of in vitro culture on somaclonal variation. The plants were regenerated from the following types of culture: twelve- and eighteen-month-old liquid culture of meristematic clumps (LMC₁₂₍₁₈₎), ten-month-old embryogenic cytokinin-dependent suspension (CDS), eighteen-month-old embryogenic cytokinin-dependent suspension in medium with modified NH⁺ ₄/NO₃ ^- ratio (CDS 1.7), twelve-month-old embryogenic auxin-dependent suspension (ADS), thirty six-month-old embryogenic auxin-dependent suspension in medium with modified NH⁺ ₄/NO₃ ^- ratio (ADS 1.7) and recurrent leaf callus regeneration (RLC) – repeated 5 times. The differences in the incidence of the following properties were observed: the ploidy of R₀ plants, the segregation of new morphological traits in R₁ and the germination ability of R₁ seeds. R₁ families with the segregation of new phenotypes were most numerous in CDS (62.5%) and LMC₁₈ (57.9%), next in CDS1.7 (35.7%), while the smallest number was found in LMC₁₂ (11.1%) and RLC (3.4%).Tetraploid and mixoploid plants occurred in ADS1.7 and ADS (100%) whereas CDS and RLC were observed to contain only tetraploids, respectively 33.3% and 55.2%. There were no changes of ploidy after LMC₁₂, LMC₁₈ and CDS1.7. Among new phenotypes there were such that have not been described so far in cucumber: ginkgolike leaf (gll), yellow-green chlorophyll mutants (y-gc), serrate margin of corolla in male and female flowers (smc). This revised version was published online in August 2006 with corrections to the Cover Date.     

Differentiation of interglycosidic linkages in permethylated flavonoid glycosides from linked-scan mass spectra (B/E)

A series of per-O-methylated flavonoid di- and tri-glycosides, linked with 1-2 and/or 1-6 glycosidic bonds between sugar rings that were isolated from different plant materials were analyzed. It was demonstrated that the fragmentation behavior of permethylated flavonoid glycosides is dependent on the glycosidic bond placement between sugars. Y(n) type fragment ions, created after glycosidic bond cleavage with oxygen retention on sugar at the reducing end for permethylated compounds, were observed in the normal and linked-scan mass spectra recorded for alpha(1-2) bonded conjugates of flavonoid di- and tri-glycosides. Moreover, for alpha (1-6) linked glycosides, Y fragments created after rearrangement and elimination of internal sugar residues were observed in addition to Y(n) type ions, but these fragment ions were not registered in normal desorption ionization spectra. This second type of fragmentation was also reported previously in collision-induced dissociation tandem mass spectrometry (CID MS/MS) spectra of some oligosaccharides and flavonoid glycosides, but their presence was independent of the glycosidic bonds placement between sugar rings.     

Effect on production and carcass quality characteristics of wether lambs of crossing Blackheaded Mutton,Ile de France and Texel rams with Polish Merino ewes

During the period 1972–1974 investigations were made on 120 wethers fattened semi-intensively to a weight of 45 kg liveweight. These lambs originated from commercial crosses of the Polish Merino with rams of the breeds Blackheaded Mutton (B), Ile de France (IF) and Texel (T) as well as pure Merino (M) in the control group.The use of rams of these meat-producing breeds as compared with Merino gave a better growth rate: an increase in the group IF × M of 6.2%, B × M of 7.8% and T × M of 8.8% respectively. The feed consumption, expressed in starch equivalents per kg of weight, decreased in group IF × M by 11.6%, B × M by 15.1% and T × M by 15.6%, and total digestible protein decreased in the group IF × M by 12.1%, B × M by 16.6% and T × M by 16.9%.Crosses with Blackheaded Mutton and Ile de France rams did not fundamentally change the tissue composition of the lamb carcasses. However, the use of Texel rams resulted in carcasses with a higher lean content and less fat as compared with pure Merino (difference 5.7%) as well as with other crosses (differences 4.2–4.8%).Rams of the meat breeds tested did not greatly influence dressing percentage and carcass yield in terms of bone content and the proportion of valuable cuts; nor were dimensions affected, except for a decrease in length of 2–3.5% compared with Merino.     

Spatial and Temporal Changes of Jet Fuel Contamination in an Unconfined Sandy Aquifer

Leakage of jet fuel from an oil skimmer has resulted in soil and groundwater contamination at the fire fighting training site of the Oslo international airport. The site is located on an unconfined sandy aquifer formed by the Gardermoen ice contact delta. A strong relationship between hydrogeological settings and spatial distribution of the hydrocarbons has been observed. Leaching hydrocarbons constitute a source of dissolved BTEX and naphthalene. Concentration of the dissolved hydrocarbons, electron acceptors and metabolic by-products has revealed concomitant changes in space and in time, in response to groundwater fluctuations. Correlation between high concentrations of hydrocarbons, elevated alkalinity and depleted electron acceptors supports the hypothesis that intrinsic bioremediation has been taking place at the site. Naphthalene, however, has been found persistent under anoxic conditions. Calculations of biodegradation potential indicates that Fe(III) reduction, methanogenesis, nitrate and sulphate reduction are the main factors controlling biodegradation.     

Historical changes in land use influence current habitat preferences of large herbivores

Landscape Ecology - Predicting habitat use patterns is a key issue in the management of large herbivore populations. Particularly, indicators providing a model of the spatial distribution of a...     

Admixture analyses and phylogeographic relationships reveal complete genetic distinctiveness of Polish farm and wild red foxes (Vulpes vulpes) and the North American origin of farm‐bred individuals

A number of studies showed that many mtDNA haplotypes were shared among contemporary farm red foxes bred on different continents and the historical wild red foxes of North American origin. Therefore, in this study, the population genetic structure and phylogeographic relationships of Polish red foxes kept on fur farms and their wild conspecifics were investigated to assess the ancestry of the farm red foxes in Poland. A total of 330 tissue samples (200 from farm foxes and 130 from wild foxes) were used for the genetic analyses. Thirty microsatellite loci and two regions of mtDNA were used to assess the level of admixture between farm‐ and wild red foxes, to construct haplotype networks and create a phylogenetic tree. The genetic structure analysis clearly indicated two genetic clusters as being the most probable number of genetically distinct populations. The fixation index revealed a significant genetic distance between the farm‐ and wild red fox populations (F_ST = 0.27, p < 0.05). Haplotype networks based on frequencies showing relationships between concatenated haplotypes of Polish farm‐ and wild red foxes and the constructed phylogenetic tree clearly indicated two genetically distinct groups. The results of this study provide strong evidence confirming the North American origin of red foxes bred on Polish farms and the genetic distinctiveness of both studied populations.     

A simple and efficient CRISPR/Cas9 platform for induction of single and multiple,heritable mutations in barley (<Emphasis Type="Italic">Hordeum vulgare</Emphasis> L.)

Background

Genome editing of monocot plants can be accomplished by using the components of the CRISPR/Cas9 (clustered regularly interspaced short palindromic repeat/CRISPR associated Cas9) technology specifically optimized for these types of plants. Here, we present the development of RNA-guided Cas9 system for simplex and multiplex genome editing in barley.

Results

We developed a set of customizable RNA-guided Cas9 binary vectors and sgRNA modules for simplex and multiplex editing in barley. To facilitate the design of RNA-guided Cas9 constructs, the pBract derived binary vectors were adapted to Gateway cloning and only one restriction enzyme was required for construction of the sgRNA. We designed a synthetic, codon optimized Cas9 gene containing the N terminal SV40 nuclear localization signal and the UBQ10 Arabidopsis 1st intron. Two different sgRNAs were constructed for simplex editing and one polycistronic tRNA-gRNA construct (PTG) for multiplex editing using an endogenous tRNA processing system. The RNA-guided Cas9 constructs were validated in transgenic barley plants produced by Agrobacterium-mediated transformation. The highest mutation rate was observed in simplex editing of the cytokinin oxidase/dehydrogenase HvCKX1 gene, where mutations at the hvckx1 locus were detected in 88% of the screened T₀ plants. We also proved the efficacy of the PTG construct in the multiplex editing of two CKX genes by obtaining 9 plants (21% of all edited plants) with mutations induced in both HvCKX1 and HvCKX3. Analysis of the T₁ lines revealed that mutations in the HvCKX1 gene were transmitted to the next generation of plants. Among 220 screened T₁ plants we identified 85 heterozygous and 28 homozygous mutants, most of them bearing frameshift mutations in the HvCKX1 gene. We also observed independent segregation of mutations and the Cas9-sgRNA T-DNA insert in several T₁ plants. Moreover, the knockout mutations of the Nud gene generated phenotype mutants with naked grains, and the phenotypic changes were identifiable in T₀ plants.

Conclusions

We demonstrated the effectiveness of an optimized RNA-guided Cas9 system that can be used for generating homozygous knockout mutants in the progeny of transgenic barely plants. This is also the first report of successful multiplex editing in barley using a tRNA processing system.