[Embryo initiation from Pinus sibirica megagametophytes in in vitro culture].

Megagametophytes of Siberian pine were cultured on an in vitro culture medium 1/2 LV supplemented with growth regulators 2,4-dichlorophenoxyacetic acid (2,4-D) and benzylaminopurine (6-BAP) to form embryos. The competency of somatic cell of explants to embryogenesis manifested itself in an organized growth and polarity. A coenocyte consisting of long vacuolated cells was formed in the megagametophyte culture. Then, the migration of the nuclei to one of the poles of the cell, their division, and formation of embryoids was observed. The megagametophyte culture of the Siberian pine differed from the zygotic embryo culture by the absence of asymmetric division in the vacuolated cell.

[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.

[Microsporogenesis and pollen formation in Scotch pine (Pinus sylvestris L.)].

The prolongation of a warm season during autumn, typical for the Siberia in recent 5-6 years, caused changes in the terms of reproductive processes in microsporangia of Scotch pine in the Krasnoyarsk region. The meiosis started in the autumn of the year, preceding the pollination, and comes to the end in next spring, whereas usually this process occurs in the spring of the year of pollination. In such conditions the division of microsporocytes accompanied with different abnormalities, causing various anomalies and the incomplete development of pollen grains.

[Patterns of somatic embryo formation in Siberian larch: embryological aspects].

Somatic embryogenesis was induced in Siberian larch by in vitro culturing zygotic embryos at different developmental stages. Cultures were grown in modified Murashige and Skoog medium supplemented with hormones 2,4-dichlorophenoxyacetic acid (2 mg/l) and 6-benzylaminopurine (0.5-1 mg/l). The success of somatic embryogenesis in this species depended on the tree genotype and developmental stage of embryos used for culturing. Somatic embryogenesis from immature zygotic embryos at the stage of cotyledon initiation was most active. After 5-10 days, such embryos formed the embryogenic tissue including two cell types--elongated highly vacuolated embryonic tubes and small embryonic cells. Somatic embryos were isolated from proliferating embryogenic tissues after 2 months of culture.

[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.

[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 the development of Siberian stone pine megagametophytes and embryos in vitro]. / Osobennosti razvitiia megagametofitov i zarodyshei sosny kedrovoi Sibirskoi v kul'ture in vitro.

Seedlings were grown in vitro from fertilized eggs and immature embryos of the Siberian stone pine. Cultivation of megagametophytes on a hormone-containing Murashige-Skoog medium from the egg formation until the globular embryo stage made it possible to manipulate fertilization and embryogenesis. Immature embryos are the most promising for in vitro cultivation. Their maturation and germination proceed within seven days of cultivation. When zygotic embryos were cultivated, adventitious buds were formed from cells at the cotyledon base and tips. When adventitious buds were subcultivated on a medium containing benzylaminopurine and naphthylacetic acid, organogenic callus and shoots were formed. Thus, cultivation of megagametophytes and embryos of the Siberian stone pine led to the completion of embryogenesis and formation of viable of seedlings.