Journal of Applied Biotechnology

Number V3N1, 2015

1-12
Growth and Biochemical Characterization of Olive (Olea europaea L., cv. Raja) Callus Cultures Differing in Proliferation Pattern

Authors: Valentina Iori, Massimo Zacchini

Abstract

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Number of views: 476

Olive is the most diffused fruit crop in the Mediterranean region, and the improvement of its productivity by conventional and biotechnological approaches is currently required. In vitro cultures have been proven to be very useful for this purpose. In this work, callus cultures of olive (Olea europaea L., cv. Raja) were established from basal tissues of nodal explants and subcultured on OM medium in a growth chamber at 25 °C with a 16h-light photoperiod. Cell clusters with different morphological aspect originated from culture, giving rise to two distinct callus lines that were isolated and long subcultured. Growth and biochemical analyses were then performed to characterize the different proliferation pattern of the two olive callus lines. Compact-type proliferating cultures showed lower growth ability, evaluated by relative growth rate (RGR), than friable-type ones. At biochemical level, differences in pigment, oxidative stress and antioxidative defense marker contents between the two callus lines were observed. In particular, higher chlorophyll and total carotenoid content, higher hydrogen peroxide content and antioxidative enzyme (ascorbate peroxidase, APX, EC 1.11.1.11; glutathione reductase, GR, EC 1.6.4.2; catalase, CAT, EC 1.11.1.6) activities were observed in compact-type proliferating cultures compared to friable ones. On the contrary, at polyamine (PA) level, a higher putrescine (Put) content was detected in the friable-type proliferating cultures. Results evidenced that proliferation of olive callus cells can be characterized by different features involving morphological aspects and biochemical traits associated to cell oxidative condition.

13-24
Herbaceous Forage Legumes Adaptation in Acidic Soils in South-Kivu, D. R. Congo

Authors: M. M. D. Katunga, J. B. B. Muhigwa

Abstract

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Number of views: 481

In D. R. Congo, agriculture is still managed in extensive system with crop integration especially in South-Kivu Province. Demographic pressure on the land is highly due to unequal distribution. The objective of this study is to test adaptation of improved herbaceous forage legumes selected in Tropical America by “Centro International de Agricultura Tropical” (CIAT). Agronomic performance and farmer participatory evaluation in using various agroecological zones in South-Kivu, were used to draw a model for similar areas. Forages were installed randomly in the blocks with 3 replicates. In spite of the acidic soils, forages recommended were, S. guianensis 11995, S. guianensis Cook, C. molle and D. intortum. The second step should be to adapt these forages in the farmers cropping systems. The seasons didn’t influence in general the yield production. All the accessions of V. unguiculata and C. ternatea did not adapt anywhere and C. brasiliensis was sensitive to diseases and insects.

25-33
Feeding Rabbits in Traditional System With Improved Forage Legumes in South-Kivu. D. R. Congo

Authors: D. M. M. Katunga, J. B. Muhigwa, J. C. K. Kashala, Y. Mbuyi, V. Okombe, K. F. Balemirwe

Abstract

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Number of views: 488

To investigate for an efficient feeding of rabbits in the traditional system essentially with some improved high yield forage legumes. Two animal nutrition trials were conducted from September 2011 to February 2012 in Mugwahi farm in Nyangezi, South-Kivu, D. R. Congo. Five female rabbits replicated three times were fed essentially in traditional system with supplements of improved forage legumes; first with Leucaena diversifolia from the selection of CIAT in Colombia and second with Desmodium intortum. Five others as local controls were fed only on local forages. The palatability evaluation showed that Calliandra calothyrsus was most appreciated by rabbits; Desmodium intortum and Leucaena diversifolia had a moderate palatability. The weight gain showed that rabbits which received a supplement of Leucaena diversifolia grew better than those fed on local forages. In term of palatability, the various forages supplied to female rabbits only C. calothyrsus performed significantly with a high RIP during the first trial and the improved forages D. intortum and L. diversifolia had a moderate one during the second trial. Regarding the weight gain of rabbits, the introduced forages; L. diversifolia performed better than the local ones. The cropping of improved legumes constitutes an alternative to avoid long walks to collect fodder which is scarce in villages. These good forages on farm could remain available even in the dry season. Studies to determine the constraints of adoption of forage crops by the farmers will improve forage production.

34-44
Application of Circular Polymerase Extension Cloning to Generate Infectious Clones of a Plant Virus

Authors: Dey K. Kishore, W. B. Borth, M. J. Melzer, John S. Hu

Abstract

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Number of views: 444

Infectious clones of viruses allow elucidation of viral gene function in plants. This study presents an adaptation and expansion of the circular polymerase extension reaction that enables both cell-based and cell-free one-step assembly of a plant viral cDNA. To demonstrate its infectious nature, the generated clone was introduced into plants by Agrobacterium infiltration. This technique eliminates the cumbersome strategy of restriction enzyme digestion and ligation and instead relies on the sequence specificity of overlapping PCR fragments to assemble a complete functional infectious clone of a virus. This technique is rapid and potentially applicable for cloning virus genes which may be difficult to clone using conventional approaches due to toxicity problems that may be encountered when cloning in E. coli.

45-62
Isolation of Geobacillus stearothermophilus L-arabinose Isomerase for the Production of the Novel Food Ingredient D-tagatose

Authors: Monika Van Holsbeeck, Efstathia Tsakali, Evelien Syryn, Carolien Van den Bussche, Guido Aerts, Jan Van Impe, Ilse Van de Voorde

Abstract

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Number of views: 498

D-tagatose, a low-calorie bulk sweetener, can be produced from D-galactose with an L-arabinose isomerase (L-AI) from Geobacillus stearothermophilus. Isolation of L-AI enzyme, intracellularly expressed in Escherichia coli, was studied by means of sonication, by chemical cell disruption and by a combined disruption approach. Conditions for cell disruption through sonication, e.g., sonication time, duty cycle, power and sample volume were determined for the wild type (WT) enzyme which was produced without an inducible expression system. The highest release was observed at a power of 120 W after 5 minutes of sonication with a duty cycle of 50% and sample volume of 60.0 mL resulting in a relative L-AI activity of 92.5 ± 0.9%. Chemical cell disruption with 16.5 mM triton X-100 for 17 hours gave even better results compared to sonication, namely, a relative L-AI activity in the crude extract of 95.6 ± 1.2%. Additional experiments were performed on cell disruption of the wild type enzyme produced with an inducible expression system (WTi). Treatment of WTi enzyme with 16.5 mM triton X-100 for 17 hours led to a lower relative L-AI activity, namely, 84.0 ± 0.5%. A combined approach of a prior chemical lysis with triton X-100 followed by sonication led to a further increase of L-AI activity towards 89.6 ± 0.3%.

63-74
Cellulase Induction in Three Aspergillus Species Isolated From Artemisia annua L. Plantation Soil Using Different Cellulose Substrates

Authors: A. I. Ogbonna, P. O. Nwadiaro, A. Chuku, C. I. C. Ogbonna, F. C. Onwuliri

Abstract

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Number of views: 496

This study was aimed at isolation and screening of fungal species associated with Artemisia annua Plantation soils from one of the under studied areas in Plateau state, Nigeria for cellulase activity. A total of thirteen fungal species were isolated from various locations within the A. annua Plantation and were screened for cellulase production. Agar plate assay was carried out using basal medium supplemented with 1% Carboxymethylcellulose (CMC) powder and staining with 0.1% Congo red solution after the incubation period. Among these species, Aspergillus fumigatus, A. niger and A. terreus were more predominant and were recorded as cellulase producing species. They have shown to possess cellulose degrading ability and exhibited maximum zones of hydrolysis on Carboxymethylcellulose medium and were selected for enzyme assay using submerged fermentation (SmF). Enzyme production was analyzed by Dinitrosalicylic acid (DNSA) methods and the enzymes assayed for were CMCase (β-1,4-endoglucanase), β-glucosidase and FPase (total cellulose) using Carboxymethylcellulose, cellulose acetate and Filter paper as substrates respectively. The highest cellulase activity was observed on the 3rd day in A. niger with enzyme production of 0.045IU/ml and 0.040IU/ml on CMC and filter paper media respectively. A. fumigatus had high enzyme activity of 0.037IU/ml and 0.025IU/ml on filter paper and cellulose acetate media respectively. Highest enzyme production of 0.034IU/ml was recorded for A. terreus on the 3rd day on cellulose acetate medium. These fungal species could be employed specially to perform in situ environmental applications involving cellulose biodegradation of wastes.