The nature of the explant, seedling age, medium type, plant growth regulators, complex extracts (casein hydrolysate, coconut milk, malt extract and yeast extract) and antioxidants (activated charcoal, ascorbic acid, citric acid and polyvinylpyrrolidone) markedly influenced in vitro propagation of Gymnema sylvestre. A maximum number of shoots (57.2) were induced from 30 day old seedling axillary node explants on Murashige and Skoog (MS) medium containing 6-benzyladenine (1 mg l−1), kinetin (0.5 mg l−1), 1-napthalene acetic acid (0.1 mg l−1), malt extract (100 mg l−1) and citric acid (100 mg l−1). High frequency of rooting was obtained in axillary node explant derived shoots (50%) on half strength MS medium supplemented with IBA (3 mg l−1). The plantlets, thus developed, were hardened and successfully established in natural soil.

In vitro micropropagation of Gymnema sylvestre - a multipurpose medicinal plant.

Efficient protocols of axillary bud multiplication and indirect organogenesis were established for Holostemma ada-kodien Schult. (Asclepiadaceae). Murashige and Skoog (MS) medium supplemented with 2.0 mg l–1 N6-benzylaminopurine (BAP) and 0.5 mg l–1 indole-3-butyric acid (IBA) induced an average of eight shoots per node and was the best for axillary bud proliferation. Subsequent cultures enhanced the number of shoots. The explant source of callus and the growth regulator inducing the callus exhibited significant influence on organogenesis. Callus developed from the basal cut end of the node explants differentiated more than 15 shoots on MS medium fortified with 1.5 mg l–1BAP. Callus from internode explants developed fewer shoots than callus from the basal cut ends of node explants. Leaf-derived callus did not undergo organogenesis. The abscission of leaves and shoot tips of the developed shoots was prevented by the addition of AgNO3 or CoCl2, but with a concomitant significant reduction in the number of shoots. Half-strength solid MS or liquid medium with 0.05 mg l–1 IBA exhibited the best in vitro rooting. Ninety percent of the rooted shoots survived in the field.

Rapid propagation of Holostemma ada-kodien Schult., a rare medicinal plant, through axillary bud multiplication and indirect organogenesis

An efficient protocol for in vitro propagation of an aromatic and medicinal herb Ocimum basilicum L. (sweet basil) through axillary shoot proliferation from nodal explants, collected from field-grown plants, is described. High frequency bud break and maximum number of axillary shoot formation was induced in the nodal explants on Murashige and Skoog (1962) medium (MS) containing N6-benzyladenine (BA). The nodal explants required the presence of BA at a higher concentration (1.0 mg·l−1, 4.4 µM) at the initial stage of bud break; however, further growth and proliferation required transfer to a medium containing BA at a relatively low concentration (0.25 mg·gl−1, 1.1 µM). Gibberellic (GA3) at 0.4 mg·l−1 (1.2 µM) added to the medium along with BA (1.0 mg·l−1, 4.4 µM) markedly enhanced the frequency of bud break. The shoot clumps that were maintained on the proliferating medium for longer durations, developed inflorescences and flowered in vitro. The shoots formed in vitro were rooted on half-strength MS supplemented with 1.0 mg·l−1 (5.0 µM) indole-3-butyric acid (IBA). Rooted plantlets were successfully acclimated in vermi-compost inside a growth chamber and eventually established in soil. All regenerated plants were identical to the donor plants with respect to vegetative and floral morphology.

In vitro clonal propagation of an aromatic medicinal herb Ocimum basilicum L. (sweet basil) by axillary shoot proliferation

Caulogenic responses of various explant types from 12-month-old plants of Hemidesmus indicus were tested Second and third visible nodes (05 cm) from the apex and root segments (05 cm) were the most and least regenerative respectively, with the formation of 937 and 26 shoots in 4 weeks on half strength MS medium supplemented with 222 μM BA and 107 μM NAA and 444 μM BA and 269 μM NAA respectively Caulogenic ability of the nodes decreased with increasing maturity Shoot buds initiated upon the young nodes on day 10 developed into 72 cm long shoots within 4 weeks thereby making a shoot elongation phase unnecessary Nodal explants of the in vitro raised shoots subcultured in the same medium produced 932 shoots of 71 cm length in 3–4 weeks, similar to those of the mature plant-derived nodes Multiplication through subculture of the nodes up to 25 passages of 4 weeks each was achieved without decline Shoot cultures were rooted in quarter salt strength MS medium containing 98 μM IBA and the rooted plants were hardened for establishment in pots at 96% rate Four months after establishment, the micropropagated plants were stable and showed uniform morphological and growth characteristic After 12 months of cultivation in the field, on an average micropropagated plant consisted of 4–5 shoots, 5–8 branches per shoot and increased root biomass (135 g) compared to the poor growth (single shoot and 2–3 branches) and root production (46 g) values obtained with plants raised from conventional rooted stem cuttings The concentration of the root specific compound, 2-hydroxy 4-methoxy benzaldehyde per plant was 2–3 fold higher in micropropagated plants though on unit dry root biomass (012% per g dry wt) basis it remained the same between two sources of plants

Micropropagation of Hemidesmus indicus for cultivation and production of 2-hydroxy 4-methoxy benzaldehyde

Protocol for rapid in vitro propagation of Ceropegia candelabrum L. (Asclepiadaceae) through axillary bud multiplication was established. Murashige and Skoog (MS) medium with 8.87 μM N
6-benzyladenine (BA) and 2.46 μM indole-3-butyric acid (IBA) was best suited for axillary bud proliferation inducing a mean of eight shoots / node. Excision and culture of the nodal segments from the in vitro shoots on fresh medium with same concentrations of BA and IBA facilitated development of more than 10 shoots / node. Subsequent cultures enhanced the rate of shoot proliferation. Shoots developed were rooted best on half strength MS with 0.49 μM IBA. Starting from a single node explant, 250 rooted shoots were obtained within 120 days. Plantlets established in pots exhibited 90% survival. Plantlets successfully established in field exhibited morphological characters identical to mother plants.

Rapid in vitro propagation of medicinally important Ceropegia candelabrum

A procedure for rapid in vitro propagation of the aromatic and medicinal plant Hemidesmus indicus (L.) R.Br. (Family Asclepiadaceae) from nodal explants is described. The highest shoot multiplication rate of 8.2 ± 0.4 shoots/explant with a 95% frequency was achieved in S weeks culture period on Murashige and Skoog medium supplemented with 1.15 μM kinetin and 0.054 μM α-naphthaleneacetic acid. Excised shoots were rooted on the same basal medium supplemented with 1.15 μM kinetin and 7.35 μM indole-3-butyric acid. Shoots derived from subcultures exhibited better rooting response than those from primary cultures. After a hardening phase of 2 weeks, there was a 70% transplantation success in the field.

J. Patnaik

Papers

Micropropagation of Hemidesmus indicus (L.) R. Br. through axillary bud culture