Genomics and synthetic biology as a viable option to intensify sustainable use of biodiversity

The Amazon basin is an area of mega-biodiversity. Different models have been proposed^1-8^ for the establishment of an effective conservation policy, increasing sustainability and adding value for biodiversity. Currently, a broad spectrum of technologies from genomics to synthetic biology is available, and these permit the collection, manipulation and effective evaluation of countless organisms, metabolic pathways and molecules that exist as potential products of a large, biodiverse ecosystem. The use of Genomics and synthetic biology may constitute an important tool and be a viable option for the prospection, evaluation and manipulation of biodiversity as advocated as well as be useful for developing methods for sustainable use and the production of novel molecules

Spinning Gland Transcriptomics from Two Main Clades of Spiders (Order: Araneae) - Insights on Their Molecular, Anatomical and Behavioral Evolution

Characterized by distinctive evolutionary adaptations, spiders provide a comprehensive system for evolutionary and developmental studies of anatomical organs, including silk and venom production. Here we performed cDNA sequencing using massively parallel sequencers (454 GS-FLX Titanium) to generate ∼80,000 reads from the spinning gland of Actinopus spp. (infraorder: Mygalomorphae) and Gasteracantha cancriformis (infraorder: Araneomorphae, Orbiculariae clade). Actinopus spp. retains primitive characteristics on web usage and presents a single undifferentiated spinning gland while the orbiculariae spiders have seven differentiated spinning glands and complex patterns of web usage. MIRA, Celera Assembler and CAP3 software were used to cluster NGS reads for each spider. CAP3 unigenes passed through a pipeline for automatic annotation, classification by biological function, and comparative transcriptomics. Genes related to spider silks were manually curated and analyzed. Although a single spidroin gene family was found in Actinopus spp., a vast repertoire of specialized spider silk proteins was encountered in orbiculariae. Astacin-like metalloproteases (meprin subfamily) were shown to be some of the most sampled unigenes and duplicated gene families in G. cancriformis since its evolutionary split from mygalomorphs. Our results confirm that the evolution of the molecular repertoire of silk proteins was accompanied by the (i) anatomical differentiation of spinning glands and (ii) behavioral complexification in the web usage. Finally, a phylogenetic tree was constructed to cluster most of the known spidroins in gene clades. This is the first large-scale, multi-organism transcriptome for spider spinning glands and a first step into a broad understanding of spider web systems biology and evolution

Application Of Electrochemically Produced Aluminum Hydroxide Gel For Prepurification Of Recombinant Synthetic Green Fluorescent Protein Produced In Tobacco Leaves.

The use of recombinant proteins has increased greatly in recent years, as also have increased the number of techniques and materials used for their production and purification. Among the different types of bioreactors being studied, there is a general consensus among scientists that production in green plant tissues such as leaves is more feasible. However, the presence of chlorophyll and phenolic compounds in plant extracts, which can precipitate and denature the proteins besides damaging separation membranes and gels, makes this technology impracticable on a commercial scale. In the present work, the adsorption to electrochemically produced aluminum hydroxide gel was applied as a prepurification step for recombinant synthetic green fluorescent protein (sGFP), also referred to as enhanced green fluorescent protein, produced in Nicotiana benthamiana leaves. Removal efficiencies of 99.7% of chlorophyll, 88.5% of phenolic compounds, and 38.5% of native proteins from the N. benthamiana extracts were achieved without removing sGFP from the extracts. As electrochemical preparation of aluminum hydroxide gel is a cost-effective technique, its use can substantially contribute to the development of future production platforms for recombinant proteins produced in green plant tissues of pharmaceutical and industrial interest.271029-3

In situ DNA transfer to chicken embryos by biolistics

Fertilized chicken eggs were bombarded with a biolistic device. Transient expression of the lacZ gene under the control of a human cytomegalovirus (CMV) promoter was assessed after in situ gene transfer using this approach. The influence of different pressures, vacuum levels and particles was tested. Survival rate improved as particle velocity decreased, but resulted in lower levels of expression. The best survival and expression were obtained with gold particles, a helium gas pressure of 600 psi and a vacuum of 600 mmHg. Under these conditions, all bombarded embryos showed <FONT FACE="Symbol">b</FONT>-galactosidase activity, indicating that this was an effective method for transformation of chicken embryos.<br>Ovos fertilizados de galinha foram bombardeados através da técnica de biobalística. A expressão transiente do gene lacZ, sob o controle do promotor humano citomegalovírus, foi verificada após a transferência in situ. Diferentes níveis de pressão de gás hélio, vácuo e tipos de partículas foram testados. A taxa de sobrevivência aumentou à medida que a velocidade das partículas diminuíram, entretanto, o nível de expressão foi menor. Os melhores resultados, combinando taxa de sobrevivência e expressão, foram obtidos com partículas de ouro, 600 libras por polegada ao quadrado de hélio e 600 mmHg de vácuo. Nestas condições, todos os embriões bombardeados apresentaram atividade da <FONT FACE="Symbol">b</FONT>-galactosidase, indicando que esta técnica é eficiente para a transformação de embriões de galinhas

Expression of functional recombinant human factor IX in milk of mice

Human factor IX is synthesized in the liver and secreted in the blood, where it participates in a group of reactions involving coagulation factors and proteins that permit sanguinary coagulation. in this work two lines of transgenic mice were developed to express the FIX gene in the mammalian glands under control of milk beta-casein promoter. the founding females secreted the FIX in their milk (3% total soluble protein). the stable integration of transgene was confirmed by southern blot analysis. the presence of the FIX recombinant protein in the milk of transgenic females was confirmed by western blot and the clotting activity was revealed in blood-clotting assays. the coagulation activity in human blood treated with recombinant FIX increased while the time of coagulation decreased. Our results confirm the production of a large amount of recombinant biologically active FIX in the mammary gland of transgenic mice.Empresa Brasileira de Pesquisa Agropecuaria (Embrapa)Hospital de Apoio de BrasiliaUniversidade Federal de São PauloConselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)International Hemophilia Training Center-World Federation of HemophiliaEmbrapa Recursos Genet & Biotecnol, BR-70770900 Brasilia, DF, BrazilUniv Brasilia, Dept Biol Celular, BR-70910900 Brasilia, DF, BrazilSGAIN, Hosp Apoio Brasilia, BR-70620000 Brasilia, DF, BrazilUniversidade Federal de São Paulo, BR-04023062 São Paulo, BrazilUniversidade Federal de São Paulo, BR-04023062 São Paulo, BrazilWeb of Scienc

Nanoscale Investigations of Synthetic Spider Silk Fibers Modified by Physical and Chemical Processes

Spider silk has biocompatibility and biodegradability properties and is known for the mechanical, physical and chemical properties that make it a promising building block in the development of novel biofibers. Its unique properties partially result from the repetitive polypeptide sequences that compose the silk proteins. The strength is related to the polyalanine motifs organized into β-sheet structures, and the elasticity is attributed to glycine-rich regions, β turns and 310 helix structures. Some alcohols were shown to induce β-sheet formation in spidroins and spider silk films, while water increases the overall structure ordering of silkworm fibroins. Furthermore, fiber stretching induces β-sheet formation in synthetic spider fibers. However, there is a lack of information relating the physical and mechanical behaviors that might contribute to improving the microstructure and performance of synthetic fibers. In this work, we reported the surface nanostructure and the nanomechanical behavior of synthetic spider fibers, which were composed of modified recombinant proteins to combine strength and extensibility motifs. Our present study evaluated synthetic fibers qualitatively and quantitatively, and indicated that atomic force microscopy (AFM) and scanning electron microscopy (SEM) were complementary tools to describe particular details of the surface structure and the mechanical features of synthetic spider fibers. Therefore, AFM and SEM would support the development of spinning systems and the characterization of novel biomaterials