Revista Colombiana de Biotecnología

6-12

Authors: Marin-Batista José, Salazar Luis, Castro Liliana, Escalante Humberto

Anaerobic digestion is an attractive technology for waste management meanwhile energy is recovering. This study evaluated the feasibility of codigesting vinasse and chicken manure (CM) as management alternative for Colombian agro industries. Biochemical methane potential was tested for different vinasse to CM ratios of 1:0, 3:1, 1:3 and 0:1 on VS basis. Vinasse and CM ratio of 3:1 increased the specific methane production up to 55% regard to the weighted specific methane production of 0.65 m3CH4/kg VS. Mixtures between the substrates had a positive synergistic effect. CM improved buffer capacity diminishing the risk on acidification by drastic pH shift during vinasse digestion. Furthermore, vinasse allowed dilution of total ammonia nitrogen concentration avoiding ammonia inhibition. Since a higher methane production, vinasse and CM co-treatment improves the energy-recovery and economic feasibility of installing biogas plant as part of the ethanol production chain. Keywords: Anaerobic codigestion, vinasse, chicken manure, biomethane potential test, synergistic effect.

24-31

Authors: Gloria Barrera, Jazmín Murcia, Jorge Cerón, Paola Cuartas, Cristian Guzmán, Laura Villamizar

The use of baculovirus as a biocontrol agent is an effective strategy, which has been gradually implemented in different production systems worldwide. For the development of a biopesticide based on baculovirus, it is necessary to have a methodology to determine the viral concentration in the process and in the finished product. In this study, a technique for viral quantification by quantitative PCR (q-PCR) was designed and optimized; therefore we used a TaqMan probe designed on granulin gene which is highly conserved. The specificity, sensitivity and reproducibility of the technique were determined. The q-PCR was able to detect and quantify isolates from the genus Betabaculovirus from five different insects species (granulovirus from Tecia solanivora, Phthorimaea operculella, Erinnys ello, Tuta absoluta and Spodoptera frugiperda) even from different geographic origins, while other isolates of baculovirus as from the genus Alphabaculovirus (nucleopolyhedrovirus from Spodoptera ornithogalli, Diatraea saccharallis or S. frugiperda) were not detected. The minimum detection limit of the technique was 6.4 x 10-4 ng /µl of DNA, equivalent to 1.25 x 103 gene copies. Additionally, intra- and inter-assays variation was minimal, demonstrating the reproducibility of technique. The applicability of the technique was evaluated for detecting granulovirus from samples of larva and soil, and to determine the virus concentration in the biopesticide formulated as emulsifiable concentrate. In conclusion, quantitative PCR was a technique reproducible, sensitive and specific to allow viral persistence studies in field, viral infection control in rearing and quality control of a biopesticide based on betabaculovirus. Key words: Granulovirus, quantitative PCR, viral persistence, quality control.

40-47

Authors: Adriana Zulay Argüello-Navarro, Niccolay Madiedo Soler, Laura Yolima Moreno-Rozo

The objective of this research was to quantify diazotrophic bacteria and compare physicochemically rhizospheric soils of three cocoa plantations (Theobroma cacao L.) in Norte de Santander Department, Colombia; for which they were characterized, differing in cultivated area, agronomic management and crop age. From serial dilutions of the samples and using the technique of Most Probable Number (MPN), In semisolid culture media (NFb, JMV, LGI, JNFb), the diazotrophs were quantified, evaluating as positive the formation of a subsurface film in the medium contained in sealed vials; equal samples were sent to the Bioambiental laboratory (UNET) for physicochemical analyzes. As a result, the evaluated samples showed deficiencies in the percentage of organic matter and elements such as Potassium, Phosphorus and Magnesium. Statistically highly significant differences in MPN were reported. The highest quantification of diazotrophs was reported in the Florilandia farm, which was characterized by drip irrigation. The highest quantification of diazotrophs was recorded in the media NFb and JMV, demonstrating a greater presence of the presumed genera Azospirillum sp. and Burkholderia sp. which are easily isolated from rhizospheric soils, unlike the genera Herbaspirillum sp. and Gluconacetobacter sp. which by their endophytic character tend to be less predominant in this type of samples. It is also concluded that the physicochemical characteristics of the soil, humidity and climatic relationships at the moment of sampling, condition the amount of root exudates and therefore are factors that conditioned the presence of diazotrophs in the samples. Keywords: Azospirillum, Burkholderia, cocoa, rhizosphere.

66-73

Authors: Paula Díaz Tatis, Carlos Andrés Zárate, Adriana Bernal Giraldo, Camilo López Carrascal

New technologies for genome edition, such as TALENs and CRISPR/Cas9 system, are a great opportunity to improve desirable features in different organisms. TALENs are the result from coupling nucleases and TALEs (Transcription Activator-Like Effectors), which are natural effectors with an important role in pathogenicity for the Xanthomonas species. Xanthomonas axonopodis pv. manihotis (Xam) is the cassava bacterial blight causal agent, and this pathogen is able to translocate its effectors to the plant cell during pathogenesis by using the type-three secretion system (TTSS). Currently, there are no standard protocols for genome edition in cassava. In this study, we explored the possibility to translocate effectors to friable embryogenic calli (FEC) through Xam inoculation, in order to establish the potential of this pathogen as a TALEN delivery system. Friable embryogenic calli derived from two different susceptible varieties (COL2215 and cv. 60444) were co-cultured with the strain Xam668 using different culture times. Subsequently, we evaluated the expression of makers corresponding to reported target genes for TALEs present in this bacterial strain. Although we were not able to demonstrate effector translocation, we established the conditions for co-culturing cassava calli and Xam and determined the effects derived from bacterial infection on embryo regeneration from FECs. Key words: Genome edition, plant tissue culture, TALEs, transformation, type-three secretion system

90-96

Authors: Jenny Paola Jiménez, Alejandro Chaparro-Giraldo

Herbicide tolerance is one of the features most used in GM crops, which has shown positive results for farmers and the environment. The starting point is the development of expression cassettes that express the characteristic of interest, they are initially constructed by standard molecular biology techniques. Currently, by bioinformatics and synthetic biology tools, it is possible to design and test the construct in silico, and then hire their synthesis. This approach allows optimizing expression by modifying the codon usage. In this work there were designed and evaluated semi-synthetic versions of genes in Nicotiana benthamiana, these genes confer tolerance to the herbicide phosphinothricin. It was made an analysis of freedom to operate in order to ensure that the designed constructs not violate intellectual property in Colombia. There were obtained two expression cassettes with freedom to operate, which express versions of the bar gene. Key words: GM Crops, freedom to operate, herbicide tolerance, codon usage.

106-111

Authors: Diana Gómez Mitjans, Vicenta Pita Bravo, Beatriz Zumalacárregui de Cárdenas

Moringa oleífera is a plant that is used as raw material in various industries, including those related to the field of chemistry such as food, pharmaceuticals and cosmetics. One of the usable parts of the tree is the seed because the content between 30 and 45% oil. They enhance its therapeutic use in the treatment of more than 300 diseases. In this research the oil extracted from the seeds of Moringa oleífera varieties of Cuban origin Supergenious, Plain and Nicaragua is characterized from solid-liquid extraction with hexane and ethanol as solvents and by the method of mechanical pressing of the species Nicaragua . Through a design of experiment 2K solute-solvent variable ratio, extraction time were analyzed, and the grain size selected runs with values greater percentage of oil extracted. Oils corresponding to the selection were characterized physic-chemical and phytochemically and were compared with varieties from different regions reported in the literature. It was shown that the pressing method is efficient, economical and has no influence on the product properties obtained. Key words: Moringa oleífera, physicochemical analysis, solid-liquid extraction, mechanical pressing, gas chromatography.

119-128

Authors: Zoraya M De Guglielmo C, Rafael Fernandez Da Silva

Full knowledge of promoters determines success in obtaining new plant cultivars through biotechnology techniques. This DNA sequence regulates the transcription of adjacent or nearby regions, which are mainly constitutive, tissue-specific or stage-specific, inducible and synthetic. This review summarizes the precise concepts, advantages and limitations of different types of promoters, including clear examples of them. Key words: Promoter, plant biotechnology, gene transcription.

140-146

Authors: Maura Isabel Díaz Lezcano, Bruno Alberto Flor Benítez, Cipriano Ramón Enciso Garay, Luis Roberto González Segnana

The objective of this work was to evaluate the effects of activated charcoal and the initial darkness conditions in the propagation in vitro of banana (Musa spp.) variety Nanicão. Were employed 64 apexes coming from shoots of approximately 20 to 30 cm. The treatments consisted in the combination of two concentrations of activated charcoal (2 and 3 g/l) and two initial dark conditions (15 and 30 days). The experimental design went completely randomized with 2 x 2 factorial arrangement and four replications. Each experimental unit consisted of four apexes. Statistical analysis consisted ANOVA with significance level of 5 %. The variables were the average rate of multiplication, survival percentage, oxidation, contamination and acclimatization of plantlets. Means were compared by Tukey test at 5 % of probability. The results showed significant difference in the interaction between the concentration of 2 g/l of activated charcoal and 15 days of initial darkness at the survival percentage from the first subculture which reached 57.30 %. In the establishment phase the percentage of contamination of the apexes was 39.06 % while that in the first subculture was 22.39 %. The oxidation in the establishment phase was 9.37 % and in the first subculture was 8.85 %. It is concluded the interaction between the concentration of 2 g/l of activated charcoal and 15 days of initial incubation period is effective in increasing survival. Key words: Musa spp, banana, survival, in vitro multiplication.