Induction of androgenic cultures in Siberian larch

Male generative buds of the Siberian larch have no organic quiescence during the autumn-winter period and are capable of completing the development of male generative structures under favorable conditions. When microsporophylls of the Siberian larch were cultivated on medium MS with 0.2 mg/l 2,4-D, embryoids of two types were obtained: directly from the microspore body and via formation of organogenic callus. This means that in the Siberian larch, direct and indirect androgenesis in vitro is possible. The Siberian larch pollen was first germinated in vitro, which was enhanced by pretreatment with hydrogen peroxide.     

Somaclonal variation of haploid in vitro tissue culture obtained from Siberian larch (Larix sibirica Ledeb.) megagametophytes for whole genome de novo sequencing

The objective of this study was to obtain a genetically stable haploid in vitro-derived line from Siberian larch (Larix sibirica Ledeb.) using megagametophyte explants, which then could be used for different molecular genetic studies, including whole genome de novo sequencing. However, cytogenetic analysis and genotyping of 11 microsatellite loci showed high levels of genomic instability and a high frequency of mutation in the obtained megagametophyte-derived callus cultures. All cultures contained new mutations in one or more microsatellite loci.     

Genetic diversity, population structure, and differentiation of Siberian larch, Gmelin larch, and Cajander larch on SSR-marker data

The genetic diversity of SSR markers was studied in six populations of Siberian larch, two popu-lations of Gmelin larch, and four populations of Cajander larch. Seven pairs of nuclear microsatellite loci were used for this analysis. 103 allelic variants were detected in 365 individuals of three species of larch. According to the AMOVA results, the variability proportion that characterizes the differences between three Larix species was twice as higher (15%) than the proportion that accounts for among-population differences within the species (6%). The differentiation of the populations of three species of larch based on SSR markers exceeded 12% (F _ST = 0.121). A significant correlation of the genetic distances with the geographic distances between populations was found (r = 0.835, P = 0.01).     

Temperature conditions of the formation of frost damages in conifer trees in the high latitudes of Western Siberia

Frost damage to the bottom of the stem at a height of 0.2 m and at the height of the position of the thermometer in the weather station (2 m) and higher in the Siberian spruce (Picea obovata Ledeb.) and Siberian larch (Larix sibirica Ledeb.) growing at the northern limits of their natural habitat were studied in order to reveal the upper threshold temperature conditions of their formation. Possible causes of differences in the distribution of frost damage in the stem of the spruce and larch are discussed.     

Phytohormone content in microstrobiles and androgenic callus of Siberian larch

Androgenesis in vitro in plants is a phenomenon of developmental switching of male generative cells, microspores, from their normal gametophytic to sporophytic pathway. We obtained androgenic callus and embryoids (embryo-like structures derived from microspores) of the conifer plant, Siberian larch (Larix sibirica Ledeb.) in the in vitro culture. The immune-enzyme analysis of the hormonal balance of larch androgenic cultures showed a substantial increase in the content of all phytohormones, especially cytokinins and ABA, as compared with initial explants (microstrobiles). This was evidently related to active cell divisions and embryoid formation. A comparison of androgenic cultures derived from trees nonifested and infested with larch gall midges (Dasineura rozhkovi Mam. et Nik.) revealed a cytokinin content increase (by two times) and an ABA content decrease (by two times), which indicates more intense growth of cultures derived from healthy trees. Phytohormone content in the androgenic callus was compared with their accumulation in the embryos of larch seeds harvested from noninfested trees. We concluded that successful growth of androgenic cultures and embryoid formation demand an additional medium supplement with auxins (not more than 0.5 mg/l).     

Distribution of extractive substances in wood of the Siberian larch (Larix sibirica Ledeb.)

A distribution of extractive substances by height on the trunk and radius of the wood of the Siberian larch (Larix sibirica Ledeb.) was investigated. The maximum flavonoid content in terms of dihydroquercetin (the main flavonoid of the larch wood) was shown to be in the butt part of the tree. In the radial direction, this parameter increased from the center to periphery of the heartwood and reached the maximum value at the sapwood boundary. The maximum content of arabinogalactan (AG) was observed in the bottom and in the top of the trunk. The distribution of the extractive substances was studied in roots of the Siberian larch for the first time.     

Characteristics of microsporogenesis in the Siberian larch growing under the conditions of technogenic load

Studies of microsporogenesis in the Siberian larch growing in Krasnoyarsk and its suburbs have shown that meiosis starts in October. Microsporocytes winter at prophase I (leptoneme, diploneme). Reduction divisions in male generative buds are resumed and terminated in spring, in March. However, in the case of thaws during the autumn-winter period, meiotic division proceeds in the larch buds and this leads to the formation of degrading tetrads and pollen. Hence, the organic quiescence is absent in the larch in winter. It was shown that in the larch growing in the city, meiosis proceed more asynchronously than in the background tree stands. An increase of chromosomal aberrations during the reduction division was noted under the conditions of technogenic pollution.     

Specific features of microsporogenesis in the Siberian larch growing under the conditions of technogenic load

Studies of microsporogenesis in the Siberian larch growing in Krasnoyarsk and its suburbs have shown that meiosis starts in October. Microsporocytes winter at prophase I (leptoneme, diploneme). Reduction divisions in male generative buds are resumed and terminated in spring, in March. However, in the case of thaws during the autumn-winter period, meiotic division proceeds in the larch buds and this leads to the formation of degrading tetrads and pollen. Hence, the organic quiescence is absent in the larch in winter. It was shown that in the larch growing in the city, meiosis proceed more asynchronously than in the background tree stands. An increase of chromosomal aberrations during the reduction division was noted under the conditions of technogenic pollution.__________Translated from Ontogenez, Vol. 36, No. 2, 2005, pp. 128–134.Original Russian Text Copyright © 2005 by Romanova, Tretyakova.     

Somatic embryogenesis in in vitro culture of three larch species

Embryogenic callus formation in different larch species from Siberia (Larix sibirica, L. gmelinii, and L. sukaczewii) was carried out on MSGm medium supplemented with growth regulators (2.4-D and BAP) and followed one and the same scheme: elongation of somatic cells and their asymmetric division with formation of initial and tube cells. The cells of embryo initial underwent sequential divisions and formed embryonic globules which caused the formation of somatic embryos. Somatic embryos became mature and germinated by addition of ABA and PEG into the medium. Long-term proliferating cell lines and regenerant plants were obtained in Sukachev larch and its hybrid with Siberian larch. The success of somatic embryogenesis depended on the genotype of the donor tree.     

Phenolic compounds of Siberian and Dahurian larch phloem

The paper presents results of the investigation of phenolic compounds of the bark of the Siberian larch (Larix sibirica Ledeb.) and the Dahurian larch (Larix gmelini (Rupr.) Rupr). The flavonoids, stilbenes, phenolic acids have been identified on the basis of chemical, chromatographic and spectral data. Quercetin-3-O-??-L-rhamnopyranoside has been found for the first time in the bark of the Larix genus. Phenolic compounds with pyrocatechol type of substitution of aromatic ring have been found to dominate in the phloem of the larch species considered.     

Induction of conifer immune responses by phytopathogenic fungus metabolites

The elicitor activity of compounds extracted from the mycelia of six species of phytopathogenic fungi was assessed from the sizes of necrotic lesions on the external surface of the living trunk phloem of five coniferous species inhabiting Siberia: Siberian larch (Larix sibirica L.), Scotch pine (Pinus sylvestris L.), Siberian spruce (Picea obovata Ledeb.), Siberian fir (Abies sibirica L.), and cedar pine (Pinus sibirica (Rupr.) Mayr.). The compounds for inoculation were extracted from the mycelium of ascomycetes imperfect, and basidium fungi; the living mycelia of these fungi were also used. The fungal extract or mycelium was placed into the hollows 7 mm in diameter in the trunk bark. Infection triggered the formation of hypersensitivity necrotic lesions in the inner bark exceeding in size those appeared after control wounding of four tree species (larch, pine, spruce, and cedar); fir was an exclusion. In experiments with tree trunks and conifer calluses, a dependence of immune response parameters (the sizes of necrotic lesions and the content of lignin and bound proanthocyanidins) on the quantity of the fungal preparation was elucidated. The largest necrotic lesions appeared after injection of 500μg of the fungal preparation into the hollow in the trunk, and its higher quantities did not increase the indices measured. The size of the necrotic lesion on the trunk bark is supposed to be used as a promising index characterizing the level of tree immunity and tolerance under various ecological conditions.     

Soil-moisture conditions indicated by field-layer plants help identify vulnerable forests in the forest-steppe of semi-arid Southern Siberia

High variability in soil-moisture conditions is typical for semi-arid forest-steppe ecosystems where precipitation varies greatly over time. Plant species that inhabit these environments integrate responses to broadly fluctuating wetness conditions. Indirect assessment of contrasting habitat wetness based on plant indicator values, species frequency, and species coverage was carried out in two sites representing the larch (Larix sibirica) and pine (Pinus sylvestris) forest-steppe communities. For the larch forest-steppe, we found that plant community composition and spatial structure depended strongly on wetness. In addition, we found that the vegetation was clearly differentiated into forest stands and steppe communities, depending on the slope aspect. There was also a strong correlation between dissimilarities of species composition and differences in habitat wetness revealed in the larch forest-steppe. In contrast, soil properties, such as gravel and stone content were found to be a key factor in the spatial distribution of plant species composition in the pine-forest-steppe communities. Indirect assessment of moisture conditions in the forest-steppe habitats, based on the field-layer plant species, was found to be preferable for indicating soil water deficits in the forest. Furthermore, as long-term observational data is often lacking, indirect assessment of the forest-steppe vegetation provides an opportunity to identify vulnerable forests at the marginal distribution. Based on indirect assessments of soil-moisture conditions, and taking into account differences in potential drought resistance between larch and pine forests, we concluded that increasing aridity will cause the replacement of Siberian larch by Scots pine in the South Siberian forest-steppe landscape. Consequently, in the future it is likely that forest-steppe typological diversity will decrease, and the semi-arid landscape may become more monotonous.     

Peculiarities and Determinants of Regeneration of Siberian Larch on the Upper Limit of Its Growth in the Urals

Quantitative and qualitative characteristics of cones and seeds and the emergence and survival of seedlings in the Siberian larch (Larix sibirica Ledeb.) have been studied in different parts of the ecotone at the upper boundary of arboreal vegetation (treeline ecotone) on Serebryanskii Kamen’ Mountain (Northern Urals) and on the hills (with a landmark 312 m above sea level) surrounding Chernaya Mountain (Polar Urals) from 2005 to 2011. We have found a decrease in the parameters of cones, number of seeds in the cones, their viability in laboratory with an increase in altitude, and differences in the number of seedlings between parts of the treeline ecotone. It is shown that the formation of Siberian larch generations on the Northern Urals occurs only after moist years. It is noted that, in the Polar Urals, Siberian larch produces a big seed crop every 2–3 years. It is proven that the seed production and mortality of seedlings in Siberian larch are influenced not only by air temperature and soil moisture, but also by wind load, snow depth, and soil temperature rapidly changing along the slope.     

The influence of the lower Ob River runoff on radial growth of trees

The influence of the Ob River runoff in its lower reaches on the radial growth of main forest-forming tree species-the Siberian spruce (Picea obovata Ledeb.), Siberian stone pine (Pinus sibirica Du Tour.), and larch (Larix sibirica Ledeb.)-was studied in the north of Western Siberia in various test sites both at the riverside and at a distance of 3 to 80 km from the Ob River floodplain. Differences in responses of the radial growth to air temperature in October and repeated frost damage of tree annual rings are observed in the Siberian spruce and Siberian stone pine depending on the distance from the river. The correlations of the radial growth of trees and frost damage with the effect of the Ob River runoff are discussed.     

Natural durability of selected larch and Scots pine heartwoods in laboratory and field tests

The aim of this study was to compare natural durability of Siberian larch heartwood grown in Siberia and Sweden as well as European larch and Scots pine heartwood grown in Sweden. The study was based on standard in- and above ground tests lasting 12 years but laboratory decay tests with white and brown rot fungi was also included. Field test results showed that Siberian larch heartwood from Siberia was the most durable among the studied heartwoods with a decay index of 60 after 12 years in Simlångsdalen (Sweden), while European larch heartwood grown in Sweden, was decayed to failure before the end of the test. Scots pine heartwood was found to perform similarly to Siberian larch from Siberia. No relationship could be established between natural durability of examined heartwoods and their water absorption behavior; however, strong correlation to the total amount of extractives was observed. Scots pine and Siberian larch heartwood from Siberia had 12.7 and 19.6% total extractives content respectively but the extractives composition differs. The study revealed also that lignin and monosaccharide content could not explain the variations in decay resistance of the studied heartwoods. No similarities in the natural durability revealed by laboratory and field tests were observed.     

Fatty acid composition of total lipids in embryogenic and nonembryogenic callus lines of larch

Fatty acid (FA) composition of total lipids in embryogenic and nonembryogenic calli of Siberian larch (Larix sibirica Ledeb.) was investigated by the method of GC-MS. We detected a high content of oleic acid in total lipids of embryogenic cell culture (32–56% by weight of total FA), which apparently depends on a high activity of acyl carrier protein (stearoyl-ACP-Δ9-desat-urase). At the same time, activity of Δ12-desaturase in the cells of embryogenic calli was considerably (2–3 times) lower than in nonembryogenic calli. We discuss a possibility that concentration of FA (oleic and linoleic acids) may be used as a marker of embryogenic potential when promising cell lines of Siberian larch are screened in the stage of early embryogenesis.     

Dynamics of structure and biological productivity of post-fire larch forests in the Northern Mongolia

Peculiarities of forming and growth of post-fire larch (Larix sibirica Ledeb.) forests at the southern range of their distribution in the Northern Mongolia were studied. Regularities of the stand structure and dynamics of biological productivity are analyzed and discussed in the paper. It has been proved that the structure of the organic mass of the post-fire herb-carex type larch tree stands at the southern border of forest vegetation distribution is closely related to their biometric indices as-age, density, and productivity. In comparison with tree stands from the other areas of forest vegetation, the total phytomass stock of larch phytocenoses appropriately increases by zonal gradient from forest-tundra border to the Transbaikalian southern taiga and northern regions of Mongolia. Regeneration of forest cover by edificator and formation of the original larch coenopopulation is a positive trend from an environmental point of view, because the Siberian larch in young and middle-age period in the area has a sufficiently high growth energy and rate of phytomass production, with more than twice exceeding mature forests in fixed carbon of the atmosphere and has positive values of carbon balance and total destruction of organic matter by the “input-output” parameters.     

Chromosomal mutations in Siberian larch (<Emphasis Type="Italic">Larix sibirica</Emphasis> Ledeb.) on Taimyr Peninsula

Chromosomal mutations were studied in the Norilsk population of Siberian larch (Larix sibirica Ledeb.) growing at the northeastern boundary of the species range affected by airborne smelter emissions. Different types of frequent chromosomal mutations have been revealed in the Norilsk population of L. sibirica and B-chromosomes have been found in this species for the first time.     

Accumulation of Phenolic Compounds in Conifer Callus Cultures in Response to Wood Blue-Stain Fungi

Callus cultures of Siberian larch (Larix sibiricaLedeb.) and Siberian spruce (Picea obovataLedeb.) were used to demonstrate the elicitor activity of two ophiostomatoid fungal species, Ceratocystis laricicolaand Ceratocystis polonica, as the pioneer settlers on larch and spruce, respectively. The extract from C. laricicolamycelium stimulated the accumulation of lignin in larch cells by 37% and that of bound proanthocyanidins by 25%. In spruce callus cultures, C. laricicolaand C. polonicaincreased the bound PA content by 25 and 46%, respectively. In the callus cultures of larch and spruce, the addition of extract of C. laricicolaincreased the concentration of p-hydroxybenzoic acid 13-fold and nearly 4-fold, respectively. The metabolic characteristics of accumulation of phenolic compounds in conifer cells are discussed.     

Molecular and Immunological Characterization of the First Allergenic Lipocalin in Hamster: THE MAJOR ALLERGEN FROM SIBERIAN HAMSTER (PHODOPUS SUNGORUS)*

The most frequent pet allergy is to cat and dog, but in recent years, it has become increasingly popular to have other pets, and the risk of exposure to new allergens is more prevalent. The list of new pets includes hamsters, and one of the most popular hamsters is the Siberian hamster (Phodopus sungorus). The aim of this study was the characterization and cloning of the major allergen from this hamster. The study of its allergenicity and cross-reactivity could improve the specific diagnosis and treatment for hamster-allergic patients. Thirteen Siberian hamster-allergic patients were recruited at the outpatient clinic. Protein extracts were prepared from the hair, urine, and salivary glands of four hamster species (European, golden, Siberian, and Roborovski). IgE-binding proteins were detected by immunoblotting and identified by mass spectrometry. The recombinant protein was produced in Escherichia coli and then purified by metal chelate affinity chromatography. The allergenic properties of the recombinant protein were tested by ELISA and immunoblotting, and biological activity was tested according to capacity for basophil activation. Three IgE-binding proteins were identified in extracts obtained from Siberian hamster hair, urine, and salivary glands. All proteins corresponded to the same protein, which was identified as a lipocalin. This lipocalin had no cross-reactivity with common and golden hamsters. The recombinant allergen was cloned and purified, showing similar IgE reactivity in vitro to Siberian hamster protein extracts. Also, the recombinant allergen was capable of producing biological activation in vivo. The major Siberian hamster allergen was cloned, and allergenic properties were characterized, providing a new tool for specific diagnosis of allergy to Siberian hamster.