Volume 2 Number 2 2008

CONTENTS AND ABSTRACTS

Xiuli Shen, Michael E. Kane (USA) Recent Advances in the Application of Plant Tissue Culture in Dieffenbachia (pp 82-91)

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Invited Review: Plant tissue culture has been shown to be a very important tool for the ornamental foliage plant industry. This is especially true for the foliage plant genus Dieffenbachia. The application of in vitro culture of Dieffenbachia has the potential to overcome some of the limitations associated with traditional methods of mass propagation, breeding and genetic manipulation. However, compared to other species, this approach has been limited in Dieffenbachia due to its recalcitrant nature in vitro. Recent advances in the application of plant tissue culture methods for the propagation and genetic manipulation of Dieffenbachia varieties are reviewed.

Danilo D. Fernando, Shiliang Zhang (USA) Characterization and Heterologous Expression of SLF, a Functional Homolog of the Floral Regulator LEAFY/FLORICAULA from Salix discolor (pp 92-99)

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Original Research Paper: This study was done to contribute to our understanding of the LFY/FLO gene activity in willow, a dioecious woody plant. SLF is the Salix discolor homolog of the LFY/FLO gene which was cloned from the reproductive buds of a male individual, clone S365. In situ hybridization revealed that SLF is strongly expressed in the inflorescence meristems, bracts, and floral meristems, but only weakly expressed in the vegetative meristems and leaf primordia. Since a genetic transformation system coupled with in vitro regeneration is currently not available for willow, Arabidopsis thaliana was used to analyze the functions of SLF. Transformed A. thaliana produced flowers more than two weeks earlier than the controls; furthermore, they produced terminal and solitary flowers instead of inflorescence branches. The phenotypes of the transgenic lines were dominant and heritable, demonstrating that SLF was functional and participated in the flowering of A. thaliana. Many of these phenotypes are being described for the first time from a LFY/FLO homolog from a dioecious plant. Complementation test showed that SLF was able to restore the wild-type phenotype of the lfy-6 mutant. This study revealed that SLF affected various aspects of floral development in transgenic A. thaliana and therefore, suggested that SLF is the functional homolog of the LFY/FLO gene.

Vinod Kumar Singh (India), Kamal Sharma (India/Nigeria) Physiological and Biochemical Changes during Flowering of Mango (Mangifera indica L.) (pp 100-105)

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Original Research Paper: Physiological and biochemical changes in terms of chlorophyll a, chlorophyll b, total chlorophyll, total sugar, reducing sugar, protein level, nitrate reductase enzyme activity, and biophysical attributes such as internal leaf temperature, internal relative humidity, diffusion resistance and rate of transpiration were measured in two types of mango cultivars i.e., regular bearing ‘Amrapali’ and irregular bearing ‘Chausa’, ‘Dashehari’ and ‘Langra’ cultivars during flower bud differentiation stage (November) and flower bud swelling stage (January). The effect of paclobutrazol (PP₃₃₃), a growth retardant having anti-gibberellin activity on these parameters, was also studied. Physiological and biochemical changes associated with the flowering of mango leaves in ‘Amrapali’ at the flower bud differentiation stage contained higher chlorophyll content, total sugar, total protein, nitrate reductase activity and higher diffusion resistance capacity as compared to ‘Chausa’, ‘Dashehari’ and ‘Langra’. High accumulation of these metabolites in the leaves of ‘Amrapali’ may possibly lead to floral induction even in the new shoot. A lower level of reducing sugar in regular bearing ‘Amrapali’ compared to irregular bearing ‘Dashehari’ at an advanced stage of the flower process (flower bud swelling stage) may indicate the capacity of this cultivar to form flower buds at a comparatively lower threshold level of reducing sugar content. The promotive effect of PP₃₃₃ on sugar and protein content can be attributed to its flower regulatory role in mango.

Mahdi Ramezani-Gask, Akhtar Shekafandeh, Hasan Salehi, Mohammad Jaafar Bahrani, Mansoor Taghvaei, Majid Jami Al-Ahmadi (Iran) A Comparison of Different Propagation Methods of Common Caper-bush (Capparis spinosa L.) as a New Horticultural Crop (pp 106-110)

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Original Research Paper: Erosion along with degraded soil and scarcity of water are serious threat to agriculture in Iran. Plants with the ability to complete their life cycle under these conditions could contribute in stemming erosion by establishing a good green conservative cover. Finding these plants may be among the priorities of eco-agriculture. Rehabilitation of these areas with these plants would require efficient propagation and successful establishment. Capparis spinosa L. (caper-bush) is commonly distributed in such areas. Therefore two experiments were conducted in Shiraz University, Iran, during 2005-2007 to investigate its sexual and vegetative propagation. Matured seeds were collected from natural vegetation and were treated with concentrated sulfuric acid and different concentrations of gibberellic acid (GA₃). For vegetative propagation tests, leafy and leafless semi-hardwood cuttings, with a length of 15 cm, were treated with various concentrations of indole-3-butyric acid (IBA) and/or naphthaleneacetic acid (NAA). Acid scarification was a necessary factor for water imbibition, but not for seed germination per se. The highest germination percentage (60.2%) was obtained by soaking seeds for 30 min in concentrated sulfuric acid followed by 90 min in a 200 mg l^-1 solution of GA₃. The highest rooting equal to 67.1 and 61.4% were obtained with 6000 and 9000 mg l^-1 IBA in leafy semi-hardwood cuttings. Our results suggest that stimulation of rooting in cuttings is a rapid and successful method for propagation of caper bush.

Mohd Faisal, Anwar Shahzad, Mohammad Anis (India) Somatic Embryogenesis and Plant Regeneration from Nodal Explants in Psoralea corylifolia L. (pp 111-113)

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Original Research Paper: The development of efficient in vitro regeneration systems are needed to facilitate the application of recombinant DNA technology to the improvement of crop germplasm. In the present study a simple, rapid and effective system to regenerate Psoralea corylifolia plants via direct somatic embryogenesis from nodal segments has been established. The embryogenic cells proliferated, formed somatic embryos, and were subsequently converted into normal plantlets under optimized culture conditions. The frequency of somatic embryogenesis was strongly influenced by the concentration of thidiazuron (TDZ) in the medium. The highest frequency (82%) of somatic embryogenesis was observed on Murashige and Skoog (MS) medium containing 16.0 µM TDZ. The somatic embryos when transferred to plant growth regulator-free MS basal medium, developed further to heart shaped, torpedo and cotyledonary stages within 2 weeks. Conversion of somatic embryos into plantlets was achieved by isolating somatic embryos with distinct cotyledons and transferring them onto half-strength MS medium containing 1.0 µM gibberellic acid (GA₃). Subsequently, the regenerated plantlets were successfully established in ex vitro condition with 90% survival. This is the first report on in vitro regeneration via direct somatic embryogenesis of P. corylifolia.

Kiranmayee Kasula, Shyam Prasad, Pavan Umate, Kranthi Gadidasu, Sadanandam Abbagani (India) Efficient TDZ and IAA-assisted Plant Regeneration from Cotyledon and Leaf Explants of Capsicum annuum L. – One-Step Protocol for Shoot Bud Differentiation and Elongation (pp 114-117)

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Original Research Paper: The present study was undertaken to investigate the possibility for developing a one-step regeneration protocol in Capsicum annuum L. For this, the effect of three different cytokinins on in vitro regeneration from cotyledon and leaf explants was studied in pepper cv. ‘G4’. The cytokinins used were 6-benzylaminopurine (BAP), thidiazuron (TDZ), and kinetin (Kn), individually or in combination with indole acetic acid (IAA). Although organogenesis could be obtained on medium supplemented with BAP or TDZ, the regenerated shoots failed to elongate on the same medium and showed an albino phenotype. TDZ stimulated shoot regeneration and this effect was significantly enhanced when combined with IAA. TDZ (9.0 µM) in combination with IAA (2.8 µM) proved to be optimal for induction of maximum number of shoots from cotyledon and leaf explants within 4 weeks on the same medium. The regenerated shoots elongated 3-4 cm on the same medium within 4 weeks from the beginning of the culture. Microshoots were rooted (after 3-4 weeks of culture on rooting medium) on MS medium fortified with IAA (5.7 µM) followed by transfer to the greenhouse with a subsequent 70% acclimatization. We conclude that TDZ in combination with IAA induces significant regeneration as well as shoot elongation from cotyledon and leaf explants in pepper cv. ‘G4’. For the first time our findings describe a single-step protocol for in vitro shoot regeneration and elongation in pepper.

Urmil J. Mehta, Sulekha Hazra (India) Induction of Somatic Embryogenesis in Tamarind (pp 118-122)

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Original Research Paper: Tamarind, being a leguminous tree species, is highly recalcitrant in nature. Very little work has been done on in vitro regeneration of tamarind. The present report describes the conditions optimized for regeneration of plants via somatic embryogenesis using immature zygotic embryos (IZEs) as the primary explants. Optimum response (17.5%) was obtained in explants cultured in Murashige and Skoog (MS) medium supplemented with dicamba at 5.0 mg l^-1. The cotyledons of IZEs turned brown and necrotic. Somatic embryos (SEs) developed either singly or in multiple from the axes. The response increased to 26.7% in explants cultured in the dark in medium gelled with phytagel in culture tubes. The protocol was tested and found to be effective for IZEs collected from five different trees although the frequency of response varied with the tree. SEs proliferated when subcultured through repetitive embryogenesis but the process was slow. Histological studies confirmed the direct and multicellular origin of the SEs from epidermal and subepidermal tissue of the apical region of the IZE. Development of SEs was restricted to the bipolar and cotyledon stage. Attempts to differentiate the SEs were futile although repetitive proliferation of the SEs and rooting was sporadically noted.

Avik Ray, Sabita Bhattacharya (India) Regeneration of Genetically Uniform Boerhaavia diffusa by Culture of Nodal Explants and Synthetic Seeds (pp 123-127)

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Original Research Paper: Multiple shoot production was achieved in Boerhaavia diffusa through axillary branching and differentiation of adventitious shoot buds at the base of nodal explants, cultured on Murashige and Skoog (MS) medium containing cytokinin alone or in combination with auxin. Application of kinetin and indole-3-acetic acid, both at 0.5 mg l^-l was most effective for shoot regeneration, resulting in an average of 3.6 shoots/explant within 30 days of culture. Re-use of nodal explants cyclically from newly regenerated microshoots increased the rate of regeneration and made the process repetitive. The shoot tips and nodal explants encapsulated with sodium alginate-polymerized MS nutrient (synthetic seeds) could be stored at 4°C for a period of 10 weeks with a 64% conversion rate. The morphologically identical plants derived from synthetic seeds and through multiple shoot formation were all genetically identical when checked by Randomly Amplified Polymorphic DNA (RAPD) analysis.

Shailendra Vyas, Rakhi Nagori, Sunil Dutta Purohit (India) Root Colonization and Growth Enhancement of Micropropagated Feronia limonia (L.) Swingle by Piriformospora indica – A Cultivable Root Endophyte (pp 128-132)

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Original Research Paper: In vitro-raisedplantlets of Feronia limonia (L.) Swingle were colonized using an endosymbiotic root fungus Piriformospora indica during in vitro rooting and their ex vitro transfer. Improved growth has been observed in terms of higher shoot and root length, internode diameter, area and number of leaves and both fresh and dry weights in plants showing an association with this fungus. More than 90% of such plants survived in the greenhouse and subsequently under a nursery shed. The study demonstrated the potential of P. indica as a biopriming agent for achieving better growth and survival of tissue culture raised plantlets.

Rajesh Kumar Suthar, Sunil Dutta Purohit (India) Root Colonization and Improved Growth Performance of Micropropagated Terminalia bellerica Roxb. Plantlets Inoculated with Piriformospora indica during ex Vitro Acclimatization (pp 133-136)

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Original Research Paper: Tissue culture raised plantlets of Terminalia bellerica, when inoculated with endophytic root fungus Piriformospora indica during their ex vitro acclimatization showed higher survival rate and improved overall growth. The fungus grew intracellularly and colonized roots of more than 80% of plantlets. Inoculated plantlets were significantly taller, had more leaves and greater total leaf area. Colonization of roots by the fungus promoted root growth and consequently caused an increase in biomass and total chlorophyll content, suggesting better uptake of nutrients.