Timir Baran Jha

In vitro propagation of cashewnut

In vitro plant propagation was developed for seedling shoot tips, leaf axils, and cotyledonary nodes of cashew, Anacardium occidentale. Factors affecting multiplication rate included age of explant source, explant type, medium composition, light requirements, and transfer frequency. Cotyledonary nodes produced more buds than other explant types. Nodes had a 90% viability when transferred daily to fresh medium containing activated charcoal for 7 d while exposed to continuous dark. Cultures were then exposed to low light illumination with weekly transfers. The phytohormone composition producing the most buds was 2.32 μM kinetin, 9.12 μM zeatin and 4.40 μM BA. The highest frequency of rooted shoots was obtained by treating shoots with the bacterium, Agrobacterium rhizogenes. Plants also were recovered by induction of roots using auxin treatment on propagated shoots.

Micropropagation of Cephaelis ipecacuanha rich.

Shoot cultures of ipecac, Cephaelis ipecacuanha Rich. were established by inoculating seedling nodal explants onto modified Murashige and Skoogs medium supplemented with 8 mg/l kinetin, 0.05 mg/l NAA and 200 mg/l adenine. Upto 12 new axillary shoots per explant were induced after 12 weeks incubation. Shoot cultures were also established by placing shoot tips on medium containing 0.1–0.25 mg/l NAA with 8 mg/l kinetin for 4 weeks and then to shoot multiplication medium for 8 weeks. The multiplication was maintained over several passages. Shoots were rooted using 2 mg/l IBA and normal plants were re-established.

Somatic embryogenesis from immature cotyledons of an elite Darjeeling tea clone

Abstract Cotyledons from immature embryos of Camellia sinensis O. Kuntze var. T-78, an elite Darjeeling tea clone, were exposed to various levels of 2,4-D, NAA, IBA, BAP and Kin (1–10 mg/1). While no embryos were formed in MS basal medium without growth regulators, the auxins tried were also inefficient in stimulating primary embryogenesis. Somatic embryos were induced directly on the cotyledonary explants in MS medium with 10 mg/1 BAP alone. Somatic embryo formation could be enhanced by addition of 0.5 mg/1 IBA and 80 mg/1 adenine to 10 mg/1 BAP. Embryo conversion was 2–3% in MS medium with or without growth regulators. Almost 20% of somatic embryos converted into whole plants following transfer to B5 medium containing 3 mg/1 BAP and 2 mg/1 IBA. Embryogenic potential was maintained for over 30 months by secondary embryogenesis through successive generation of embryos. Somatic embryo derived plants were successfully transferred to potted soil at 70% frequency under green house conditions. Morphologically and cytologically (2n = 30) plants are true to type.

Production of emetine and cephaeline from cell suspension and excised root cultures of Cephaelis ipecacuanha

Abstract Production of the ipecac alkaloids, emetine and cephaeline was studied in cell suspension and excised root cultures of Cephaelis ipecacuanha . A two-stage cell suspension culture was developed for enhanced accumulation of the alkaloids. In the first-stage, suspension cultures were established in Murashige and Skoogs (MS) medium containing 2,4-D and NAA which was suitable for cell growth and the second-stage culture system was composed of MS medium containing IBA, IAA and 6% sucrose which favoured alkaloid production. The production of emetine and cephaeline was greatly increased in the two-stage culture method compared to the single-stage culture. Optimal alkaloid synthesis was obtained in excised root culture of the plant in medium composed of half-strength MS salts, IBA (0.25 mgl −1 ) and 2% sucrose. A discernible higher accumulation of cephaeline in two-stage cell suspension culture as well as in excised root culture in comparison to that of the three-year-old roots was a

Effect of different hormones on Vigna tissue culture and its chromosomal behaviour

Abstract Vigna sinensis cv. 152 hypocotyl segments were grown in vitro in the presence of different auxins and cytokinins. The growth pattern was observed at different combinations of hormone. Indoleacetic acid (IAA) was most potent in root initiation but least so in callus induction. Both naphthyl acetic acid (NAA) and 2,4-dichlorophenoxy acetic acid (2,4-D) were almost equally potent in callus initiation but NAA produce healthier calli and caused some root initiation. 2,4-D could not cause any root initiation. Cytological analysis of calli revealed that 2,4-D induces more numerical chromosomal anomalies in Vigna sinensisthan NAA.

In vitro culture of Cuminum cyminum regeneration of flowering shoots from calli of hypocotyl and leaf explants

Calli from hypocotyl explant of Cuminum cyminum L. (Cumin) grew rapidly on Gamborgs B5 basal medium with the following supplements, (i) 0.5 mg/l — 2,4-D (ii) 4 mg/l — NAA plus 2 mg/l — Kinetin and (iii) 0.2 mg/l — NAA plus 0.2 mg/l — BAP, whereas calli from leaf explant in these media grew slowly. Hypocotyl and leaf calli produced roots when transferred to basal medium only and shoots in basal medium with 0.5 mg/l NAA and 0.1 mg/l BAP. Ninety percent of the shoots produced roots when they were transferred to half strength MS inorganic salts supplemented with 0.5 mg/l each of IBA and NAA.Fifty to sixty percent of rootless as well as rooted shoots produced terminal umbellate flowers on this medium.

Organogenesis and regeneration from pigmented callus in Camellia sinensis (L.) O. Kuntze cv. Nandadevi, an elite Darjeeling tea clone

Abstract Organogenesis and regeneration of adventitious shoots from stem callus in Camellia sinensis ev. Nandadevi was affected by explant type, nature of calli lines established and cytokinin type used. Callus cultures were established from three types of explants viz., stem epidermal layers (E1), transversely cut stem segments (E2) and longitudinally cut stripped stem segments (E3), cultured on half strength Murashige and Skoogs media containing 3 mg/l benzyladenine (BA), 3 mg/l BA, 50 mg/l adenine sulphate (M2). 3 mg/l Zn and 50 mg/l adenine sulphate (M3). Two calli lines were established from the three types of explants in M1, M2 and M3 medium i.e., friable green (FG) or compact green (CG), after 3 weeks of culture. Regeneration was achieved only in CG calli derived from E2 and E3 but not E1 explants, when such calli turned heterogenous with respect to pigmentation on subculture to basal medium containing BA (1–10 mg/l). Callus derived from E2 explants showed a higher regeneration frequency (90%) in basal medium with 5 mg/l BA. The maximum number of shoot buds obtained was 22.5±0.5 g calli within 4 weeks of culture. Shoots proliferated on basal medium containing 1 mg/l BA and 5 mg/l GA3. 37.2 ± 0.3 microshoots/g calli could be excised after 4 weeks and microshoots (3–4 cms) rooted on 1 4 basal media containing 6 mg/l lBA.

Searching chromosomal landmarks in Indian lentils through EMA-based Giemsa staining method

Lentil is one of the oldest protein-rich food crop with only one cultivated and six wild species. India is one important cultivator, producer and consumer of lentils and possesses a large number of germplasms. All species of lentil show 2n = 14 chromosomes. The primary objective of the present paper is to search chromosomal landmarks through enzymatic maceration and air drying (EMA)-based Giemsa staining method in five Indian lentil species not reported elsewhere at a time. Additionally, gametic chromosome analysis, tendril formation and seed morphology have been studied to ascertain interspecific relationships in lentils. Chromosome analysis in Lens culinaris, Lens orientalis and Lens odemensis revealed that they contain intercalary sat chromosome and similar karyotypic formula, while Lens nigricans and Lens lamottei showed presence of terminal sat chromosomes not reported earlier. This distinct morphological feature in L. nigricans and L. lamottei may be considered as chromosomal landmark. Meiotic analysis showed n = 7 bivalents in L. culinaris, L. nigricans and L. lamottei. No tendril formation was observed in L. culinaris, L. orientalis and L. odemensis while L. nigricans and L. lamottei developed very prominent tendrils. Based on chromosomal analysis, tendril formation and seed morphology, the five lentil species can be separated into two distinct groups. The outcome of this research may enrich conventional and biotechnological breeding programmes in lentil and may facilitate an easy and alternative method for identification of interspecific hybrids.

Indian Swertia from Eastern Himalaya: Strategies of Conservation and Biotechnological Improvements

The Eastern Indian area of the Himalaya is highly acclaimed for its rich bioresources and traditional repository of medicinal plants. This region is considered to be the natural habitat of many species and populations of Swertia, an important and diverse genus of the Gentianaceae. The genus Swertia, popularly known as chirata, is one of the most important indigenous medicinal plants of India. Habitat destruction, other human-caused stresses, and impacts of climate change have resulted in considerable loss of genetic diversity and necessitate reassessment, documentation, molecular and biochemical screening, and well-formulated biotechnological strategies for sustainable use and improvement of these high-value medicinal crops. Swertia chirata is considered the prized Indian species because of its well-characterized bioactive molecules known as iridoid and secoiridoid glycosides, xanthones and xanthone derivatives, which are effective against many conventional and unconventional ailments. The increasing demand of chirata in national and international markets, the paucity of agricultural practices, and correct exomorphological and molecular screening have resulted in intentional and unintentional adulteration. The prized species, Swertia chirata, has been labeled as being critically endangered and most of the other important species may face similar threats in the near future. The present review discusses the status of Swertia species in the Eastern Himalayas, together with biotechnological approaches for their conservation and future improvement.

Rhizogenesis fromNigella sativa protoplasts

SummaryProtoplasts were isolated for the first time from cell suspensions ofNigella sativa. These were then cultured in media and observed at regular intervals. Different concentrations of auxin and kinetin were tried with success to obtain root from the callus tissues of the protoplasts.