ISSN 10623604, Russian Journal of Developmental Biology, 2015, Vol. 46, No. 5, pp. 276–285. © Pleiades Publishing, Inc., 2015.
Original Russian Text © I.N. Tret’yakova, D.N. Shuvaev, 2015, published in Ontogenez, 2015, Vol. 46, No. 5, pp. 327–337.
Siberian dwarf pine (
belonging to the fiveneedle pines, grows in northeast
ern Siberia and occupies a vast area extending from the
northern Cisbaikalia to the Kamchatka Peninsula.
This species is of great ecological and economic value.
Its thickets play an important phytocoenotic and land
seeds are an important food
source for animals. However, this species is seriously
threatened on a large part of its range due to longterm
forest fires, which hamper its natural regeneration.
The problems of restoration of the
lation and conservation of its gene pool can be solved
by micropropagation through somatic embryogenesis.
This method makes it possible to perform mass repro
duction of this species with traits valuable for breeding
in an in vitro culture using DNA microarrays, trans
formation, and cryopreservation of derived embryo
genic lines to create a bank of valuable genotypes. The
phenomenon of somatic embryogenesis of
can serve not only for practical purposes but is also a
model system for studying the patterns of cell differen
tiation and realization of the morphogenetic develop
mental program in early ontogeny (Lelu, 1994; Von
Arnold et al., 2002; Park et al., 2006).
To date, embryogenic callus and somatic embryos
have been obtained for 28 species of the genus
(Laine and David, 1990; Salaijva et al., 1995; Garin
et al., 1998; Lelu et al., 1999; Arya et al., 2000; Kli
maszewska and Cyr, 2002; Pullman and Bucalo, 2010;
Tret’yakova et al., 2014). For more than half of them,
the culture conditions ensuring the regeneration of
plants from somatic embryos were optimized. The fre
quency of initiation of the embryogenic callus in the
majority of studied species of the genus
, it ranged from 2 to 25% (MacKay et al.,
2006), from 52.9% (Finer et al., 1989) to 76% (Kli
maszewska et al., 2001) in
, from 0.5 to 7.2%
(Carneros et al., 2009), and within 15% in
(Borcetche and Paques, 1995). Among the
fiveneedle pines, somatic embryogenesis induction
was described for
(Bozhkov et al.,
(Tret’yakova et al., 2014; Tret’yakova
and Voroshilova, 2014), and
(Noskova et al.,
2012; Tretiakova et al., 2012). The frequency of initia
tion of embryogenic callus in
was very low and did not exceed 0.2–0.5% (Noskova
et al., 2012, Tret’yakova et al., 2014).
This article presents the results of the biotechnol
ogy for deriving proliferating embryogenic cell lines of
and the analysis of productivity of the
derived embryogenic cultures during longterm culti
vation in vitro.
MATERIALS AND METHODS
The study was performed with seeds of 80–100year
trees growing in the wild in the vicinity of
the Novaya Chara village (Udokan Ridge spurs, south
E, 807 m above
sea level), near Lake Leprindo (
Somatic Embryogenesis in
of Embryogenic Lines during LongTerm Cultivation In Vitro
I. N. Tret’yakova and D. N. Shuvaev
Sukachev Institute of Forestry, Siberian Branch, Russian Academy of Sciences,
Akademgorodok 50, bld. 28, Krasnoyarsk, 660036 Russia
Received May 7, 2014; in final form, April 27, 2015
—Zygotic embryos and megagametophytes of
for cultivation in vitro were transferred
in 1/2 LV medium supplemented with growth regulators 2,4dichlorophenoxyacetic acid (2,4D) and benzy
laminopurine (6BAP) to induce somatic embryogenesis. Four stably proliferating cell lines from two geno
types were derived. The cell lines differed in the number of globular somatic embryos and the weight of
embryogenic calli. Cells of these lines were multiplied as a result of somatic polyembryogenesis via cleavage.
In the nutrient medium for maturation, mature somatic embryos were obtained. However, somatic embryos
of not all embryogenic cell lines reached maturation. In this study, plantlets were obtained in an in vitro cul
ture for the first time.
, zygotic embryos, megagametophytes, somatic embryos, proliferating embryogenic
lines, in vitro