Leela George

Plant regeneration from leaf base callus of turmeric and random amplified polymorphic DNA analysis of regenerated plants

Callus cultures were initiated from leaf bases of turmeric on Murashige and Skoogs basal medium (MS) supplemented with dicamba, picloram (2 mg l−1) or 1-naphthaleneacetic acid (NAA) (5 mg l−1) in combination with benzyladenine (BA) (0.5 mg l−1). On transfer of callus cultures to medium supplemented with benzyladenine (BA) (5 mg l−1) in combination with triiodebenzoic acid (TIBA) or 2,4-dichlorophenoxyacetic acid (2,4-D) (0.1 mg l−1), green shoot primordia were seen to differentiate from the surface of the callus. On transfer of regenerating cultures to half MS media supplemented with Kn, shoot primordia developed into well developed shoots. When shoots were transferred to medium devoid of phytohormones, complete rooted plants were obtained. Ninety percent of the plants survived to maturity on transfer to soil. Random Amplified Polymorphic DNA (RAPD) analysis of eight regenerated plants using 14 primers when separated on non-denaturing polyacrylamide gels showed 38 novel bands. About 51 bands present in the control were absent in the regenerants. The result indicates that variation at DNA level has occurred during in vitro culture.

Influence of phytohormones, carbohydrates, aminoacids, growth supplements and antibiotics on somatic embryogenesis and plant differentiation in finger millet

Cultured caryopses of finger millet (Eleusine coracana GAERTN) produced callus from shoot apices or mesocotyls depending upon the concentration of picloram and combination of cytokinins in MS basal medium. On subsequent subcultures, numerous somatic embryos differentiated from the callus on MS medium supplemented with picloram and kinetin. The embryos germinated into complete plants on medium devoid of phytohormones. When different carbohydrates were tested, basal medium containing glucose and sucrose produced the highest frequency of germinating somatic embryos. Supplementation of MS basal medium with a variety of aminoacids, osmotic agents and growth supplements had an adverse effect on the germination of embryos. Incorporation of different antibiotics such as carbenicillin, cefotaxime and streptomycin sulfate enhanced plant differentiation from somatic embryos. Cytological analysis of regenerated plants showed normal diploid chromosome number in their root tips.

Plant regeneration from peduncle segments of oil seed Brassica species: Influence of silver nitrate and silver thiosulfate

Cultured peduncle segments of B. juncea, B. campestris, B. napus, B. nigra and B. carinata produced shoot buds on Murashige and Skoog medium supplemented with benzyladenine and 1-naphthalene acetic acid. Supplementation of the media with 30 µm silver nitrate or silver thiosulfate enhanced the frequency of shoot regeneration. The regenerated shoots could be rooted at a frequency of 95% and transferred to soil where 75% survived and set seed.

Enhanced plant regeneration in pearl millet (Pennisetum americanum) by ethylene inhibitors and cefotaxime

Cefotaxime, a cephalosporin antibiotic, and different ethylene inhibitors, such as silver nitrate, cobalt chloride, nickel chloride and O-acetyl salicylic acid, significantly delayed the loss of regeneration potential in embryogenic cultures of Pennisetum americanum. In the presence of these chemicals, ethylene content in the atmosphere of the culture vessel was less than that of the control. Cefotaxime, silver nitrate and O-acetyl salicyclic acid did not have any effect on callus growth based on fresh weight, while growth based on dry weight was enhanced by O-acetyl salicyclic acid.

Somatic embryogenesis in peanut: Influence of growth regulators and sugars

Somatic embryogenesis and plant regeneration were induced from immature embryonal axes and immature cotyledons of peanut (Arachis hypogaea L. fastigata type cv JLM-1). Influence of different auxins, cytokinins and sugars on somatic embryogenesis from immature cotyledon explants was also investigated. Among the different auxins tested, 2,4-dichlorophenoxyacetic acid (2,4-d) was most effective, producing the highest frequency of responding cultures and highest average number of somatic embryos per responding culture, while dicamba, picloram, indolepropionic acid, α-naphthaleneacetic acid, 2,4,4-trichlorophenoxypropionic acid and α-naphthoxyacetic acid were also effective for embryogenesis. Indolebutyric acid, indoleacetic acid, p-chlorophenoxyacetic acid and trichlorophenoxyacetic acid were not beneficial. Among the four cytokinins tested, zeatin slightly enhanced the frequency of somatic embryogenesis, while kinetin, 6-γ-γ-dimethylallylaminopurine and benzyladenine were relatively inhibitory. Among the different carbon sources tested, sucrose was the best for embryo induction and at 6% sucrose the highest frequency of responding cultures and average number of somatic embryos per explant were obtained. For inducing embryogenesis from embryonal axes, 2,4-d was more effective than picloram. Highest plant conversion frequency from somatic embryos was obtained in presence of dicamba or NAA and using cotyledon explants.

Agrobacterium tumefaciens mediated gene transfer in peanut (Arachis hypogaea L.)

Transgenic peanut plants were produced using Agrobacterium mediated gene transfer. Primary leaf explants of peanut were co-cultivated with Agrobacterium tumefaciens LBA 4404 harbouring the binary plasmid pBI 121 (conferring β-glucuronidase activity and resistance to kanamycin) and cultured on regeneration medium supplemented with kanamycin to select putatively transformed shoots. They were rooted and plants were transferred to soil. Stable integration and expression of the transgenes were confirmed by NPT II assay, Southern blot hybridization and GUS assay.

Organogenesis and embryogenesis from diverse explants in pigeonpea (Cajanus cajan L.)

Seed and seedling expiants of pigeonpea were evaluated for organogenesis and somatic embryogenesis. De novo plant regeneration through organogenesis was obtained from mature cotyledons, primary leaves and roots of seedlings. Production of multiple shoots from the cotyledonary node was observed in cultures of whole seeds on 6-benzylaminopurine enriched medium. Somatic embryos were induced from immature cotyledons and embryonal axes, however, well-developed plants could not be derived from these embryos. The regenerants obtained through organogenesis were transferred to the field and grown to maturity.

Plant regeneration from leaf discs of peanut and pigeonpea: Influence of benzyladenine, indoleacetic acid and indoleacetic acid-amino acid conjugates

The influence of indoleacetic acid (IAA) and some IAA-amino acid conjugates in combination with benzyladenine (BA) on in vitro shoot regeneration from leaf discs of peanut and pigeonpea was investigated. The frequency of shoot regeneration and average number of shoot buds produced was dependent on the type of auxin present in the medium. Highest frequency of plant regeneration in peanut was induced by BA in combination with IAA or IAA-l-alanine, while in pigeonpea BA in combination with IAA or IAA-l-aspartic acid produced best results. Four hundred plants of peanut and 25 plants of pigeonpea were transplanted to soil.

In vitro induction of pollen embryos and plantlets in Brassica juncea through anther culture

Abstract Pollen embryogeny was induced in cultured anthers of Brassica juncea var. TM-4. A combination of cold pretreatment and high temperature shock was essential in inducing embryogenesis. Pollen grains developed directly into embryos on a high sucrose medium. However, further growth of the embryos was not normal and resulted in abnormal plantlets. From the abnormal plantlets obtained in vitro, hypocotyl explants were excised and were induced to regenerate numerous shoot buds on a medium supplemented with NAA and BA. The shoot buds developed into normal plants. Among the population of pollen plants both haploids and diploids were observed.

Direct regeneration of shoots from immature inflorescence cultures of turmeric

Plant regeneration from cultured immature inflorescence of Curcuma longa was obtained by direct shoot development on Murashige and Skoogs basal medium supplemented with BA (5 or 10 mg l−1) in combination with 2,4-D (0.2 mg l−1) or NAA (0.1 mg l−1) and TDZ (1 or 2 mg l−1) in combination with IAA (0.1 mg l−1). Regenerated shoots were grown on MS medium for further development and later transferred to medium supplemented with 0.1 mg l−1 NAA for induction of roots. Complete plants thus obtained were transferred to sterilized soil in paper cups for 3–4 weeks and then to field, where 95% of the plants survived to maturity.