Effect of calcium, BAP and putrescine on somatic embryo induction in juvenile explants of Eucalyptus grandis × E. urophylla hybrids

Considering the constant improvement of Eucalyptus cloning and the search for new technologies to produce plantlets of this species, somatic embryogenesis has attracted interest from research groups and forestry companies that use advanced genetic breeding and cloning programs. The objective of the present study was to verify the effect of concentrations and sources of calcium, concentrations and effect time of cytokinin BAP and polyamine putrescine on the induction and development of somatic embryos in juvenile explants of Eucalyptus grandis x E. urophylla. Cotyledonary explants were inoculated into culture medium containing calcium chloride (MS medium) or calcium nitrate (JADS medium) as source of calcium. Different concentrations of calcium were also used, for MS medium containing: 4.40 gL-1 (control-Ca), 6.60 gL-1 (50% increase over control - Ca 50) and 8.80 gL-1 (increase of 100% over control – Ca 100) of calcium nitrate; and for JADS medium containing: 11.81 gL-1 (Ca), 17.72 gL-1 (Ca50) and 23.62 gL-1 (Ca100) of calcium chloride. Cotyledon explants were inoculated into the primary induction medium (PIM) containing 20.71 μM picloram as growth regulator. At 10, 20 and 30 days of primary induction, the explants were transferred to the secondary induction medium (SIM) containing 20.71 μM picloram and 11.10 μM BAP or 28.36 μM putrescine. The culture medium containing calcium nitrate provided higher callogenesis when compared to the medium containing calcium chloride. The increase in calcium concentration in the media did not provide higher percentage of induction of somatic pro-embryos. However, the addition of 28.36 μM putrescine to the culture medium provided a higher percentage of induction of somatic embryogenesis. The number of somatic pro-embryos formed per explant was higher when BAP and putrescine were added to the culture medium when compared to medium containing only picloram. To obtain a greater number of somatic pro-embryos of Eucalyptus grandis × E. urophylla, the JADS culture medium containing 28.36 μM putrecine should be used.

PROPAGATION STRATEGIES OF TRIBULUS TERRESTRIS L. A PORTENTOUS MEDICINAL PLANT EMPLOYING TISSUE CULTURE TECHNOLOGY

A protocol for micropropagation of Tribulus terrestris, an important medicinal herb was established using juvenile explants viz., leaf, node and internode. All the explants were tested for callus induction on Murashige and Skoog’s (MS) medium, supplemented with BAP, NAA and 2,4-D. Among the three explants leaf explant responded well (98%) for the callus induction in the MS medium composted with BAP and NAA (4.0 and 0.5 mg/L) followed by the nodal segments (58.75%) in the same medium. Maximum number of shoot induction from the callus of leaf derived explants (91.1%) was perceived on MS medium fortified with BAP 4.0 mg/L and NAA (0.5 mg/L). Moreover, root elongation and profuse rooting percentage (77.19%) were achieved when the well-grown shoots were cultured on MS media supplemented with IAA (2.0 mg/L) for leaf callus derived shoots. The regenerated plantlets were hardened and established at 80% survival rate in hardening media encompassed with red soil, sand and vermicompost in the ratio of 1:1:1 by volume.

Micro- and Cutting Propagation of Silver Maple. I. Results with Adult and Juvenile Propagules

Clonal micropropagation studies with silver maple (Acer saccharinum L.) included experiments with various shoot. explant types, cytokinins, and stock plant maturation levels. These trials led to successful explant establishment, axillary shoot proliferation, rooting of microshoots, and establishment of plantlets in the greenhouse. Overall, the best cytokinin tested was the phenylurea derivative TDZ. Shoot proliferation on juvenile explants was poor with kinetin, 2iP, and BA. Only zeatin at 10 μm was comparable to TDZ. TDZ at 10 nm was optimal for both juvenile and adult nodal explants. Juvenile explants that were held in vitro for 4 months commonly had at least 60 axillary shoots that could be subculture or excised for rooting. Microshoots rooted within 2 weeks. Following rooting, silver maple plantlets could be transplanted into a growing medium and placed directly onto a greenhouse bench. Studies were also conducted on rooting stem cuttings (macropropagation). Single nodes from juvenile plants rooted under intermittent mist, regardless of auxin application; however, shoot-tip cuttings from adult trees rooted best when auxin in ethanol solution was applied. Chemical names used: N- phenyl- N' -1,2,3 -thiadiazol-5-ylurea (thidiazuron, TDZ), N- (2-furanylmethyl)-1H-purin-6-amine (kinetin), isopentenyladenine (2iP), benzyladenine (BA), (E)-2-methyl-4-(1H-purin-6-ylamino)-2-buten-1-ol (zeatin).