Specific tracking of xylan using fluorescent-tagged carbohydrate-binding module 15 as molecular probe

Additional file 5: Figure S3. Low-resolution XPS spectrum of UBKP surface. UBKP: unbleached kraft pulp. Unextracted pulp samples were analysed

Rapid and efficient colony-PCR for high throughput screening of genetically transformed chlamydomonas reinhardtii

Microalgae biotechnologies are rapidly developing into new commercial settings. Several high value products already exist on the market, and biotechnological development is focused on genetic engineering of microalgae to open up future economic opportunities for food, fuel and pharmacological production. Colony-polymerase chain reaction (colony-PCR or cPCR) is a critical method for screening genetically transformed microalgae cells. However, the ability to rapidly screen thousands of transformants using the current colony-PCR method, becomes a very laborious and time-consuming process. Herein, the non-homologous transformation of Chlamydomonas reinhardtii using the electroporation and glass beads methods generated more than seven thousand transformants. In order to manage this impressive number of clones efficiently, we developed a high-throughput screening (HTS) cPCR method to rapidly maximize the detection and selection of positively transformed clones. For this, we optimized the Chlamydomonas transformed cell layout on the culture media to improve genomic DNA extraction and cPCR in 96-well plate. The application of this optimized HTS cPCR method offers a rapid, less expensive and reliable method for the detection and selection of microalgae transformants. Our method, which saves up to 80% of the experimental time, holds promise for evaluating genetically transformed cells and selection for microalgae-based biotechnological applications such as synthetic biology and metabolic engineering

Red light variation an effective alternative to regulate biomass and lipid profiles in Phaeodactylum tricornutum

Abstract: Marine water diatom Phaeodactylum tricornutum is a photosynthetic organism that is known to respond to the changing light environment and adapt to different temperatures to prevent photoinhibition and maintain its metabolic functions. The objective of the present study was to test whether light shift variations in different growth phases impact the growth and lipid metabolism of P. tricornutum. Thus, we investigated R exposure in different growth phases to find the most effective light shift condition. The results showed that substituting white light (W) by red light (R) under autotrophic conditions, a condition called red shift (RS), increased biomass and lipid content compared to levels found under continuous W or R exposure alone. We observed an increase by 2-fold biomass and 2.3-fold lipid content in RS as compared to W. No significant change was observed in the morphology of lipid droplets, but the fatty acid (FA) composition was altered. Specifically, polyunsaturated FAs were increased, whereas monounsaturated FAs decreased in P. tricornutum grown in RS compared to W control. Therefore, we propose that a light shift during the beginning of the stationary phase is a low-cost cultivation strategy to boost the total biomass and lipids in P. tricornutum

Diatoms biotechnology: various industrial applications for a greener tomorrow

The benefits of the complex microscopic and industrially important group of microalgae such as diatoms is not hidden and have lately surprised the scientific community with their industrial potential. The ability to survive in harsh conditions and the presence of different pore structures and defined cell walls have made diatoms ideal cell machinery to produce a variety of industrial products. The prospect of using a diatom cell for industrial application has increased significantly in synch with the advances in microscopy, metabarcoding, analytical and genetic tools. Furthermore, it is well noted that the approach of industry and academia to the use of genetic tools has changed significantly, resulting in a well-defined characterization of various molecular components of diatoms. It is possible to conduct the primary culturing, harvesting, and further downstream processing of diatom culture in a cost-effective manner. Diatoms hold all the qualities to become the alternative raw material for pharmaceutical, nanotechnology, and energy sources leading to a sustainable economy. In this review, an attempt has been made to gather important progress in the different industrial applications of diatoms such as biotechnology, biomedical, nanotechnology, and environmental technologies

Factors associated with adverse reactions induced by caprylic acid-fractionated whole IgG preparations: comparison between horse, sheep and camel IgGs

Caprylic acid purification of IgG, currently used in the manufacture of horse-derived antivenoms, was successfully adapted for the preparation of sheep and camel IgG. Sheep IgG had a molecular mass of approximately 150 kDa, whereas camel IgG presented two bands of molecular masses of approximately 160 and 100 kDa, the latter corresponding to heavy-chain IgG, which is devoid of light chains. Horse, sheep and camel IgGs were compared by several parameters aiming at predicting their potential for induction of early and late adverse reactions. Horse and sheep IgGs showed a higher anticomplementary activity than camel IgG, and also elicited a higher anti-IgG response than camel IgG, when injected in mice. Horse IgG agglutinated human type O+ erythrocytes, whereas no such reactivity was observed in sheep and camel IgG preparations. A novel procedure was used for the detection of antibodies in human serum against animal IgGs. It was found that a pool of human sera collected in Costa Rica had a higher titer of antibodies directed against horse and sheep IgGs than against camel IgG. Overall, camel IgG showed the lowest potential for the induction of adverse reactions among the three IgGs tested.Universidad de Costa Rica/[741-A4-503]/UCR/Costa Rica/[883.701-3]/UNESCO/Costa RicaUCR::Vicerrectoría de Investigación::Unidades de Investigación::Ciencias de la Salud::Instituto Clodomiro Picado (ICP

Single domain antibodies: promising experimental and therapeutic tools in infection and immunity

Antibodies are important tools for experimental research and medical applications. Most antibodies are composed of two heavy and two light chains. Both chains contribute to the antigen-binding site which is usually flat or concave. In addition to these conventional antibodies, llamas, other camelids, and sharks also produce antibodies composed only of heavy chains. The antigen-binding site of these unusual heavy chain antibodies (hcAbs) is formed only by a single domain, designated VHH in camelid hcAbs and VNAR in shark hcAbs. VHH and VNAR are easily produced as recombinant proteins, designated single domain antibodies (sdAbs) or nanobodies. The CDR3 region of these sdAbs possesses the extraordinary capacity to form long fingerlike extensions that can extend into cavities on antigens, e.g., the active site crevice of enzymes. Other advantageous features of nanobodies include their small size, high solubility, thermal stability, refolding capacity, and good tissue penetration in vivo. Here we review the results of several recent proof-of-principle studies that open the exciting perspective of using sdAbs for modulating immune functions and for targeting toxins and microbes

Genome Editing by CRISPR-Cas: A Game Change in the Genetic Manipulation of Chlamydomonas

Microalgae are promising photosynthetic unicellular eukaryotes among the most abundant on the planet and are considered as alternative sustainable resources for various industrial applications. Chlamydomonas is an emerging model for microalgae to be manipulated by multiple biotechnological tools in order to produce high-value bioproducts such as biofuels, bioactive peptides, pigments, nutraceuticals, and medicines. Specifically, Chlamydomonas reinhardtii has become a subject of different genetic-editing techniques adapted to modulate the production of microalgal metabolites. The main nuclear genome-editing tools available today include zinc finger nucleases (ZFNs), transcriptional activator-like effector nucleases (TALENs), and more recently discovered the clustered regularly interspaced short palindromic repeats (CRISPR)-CRISPR associated protein (Cas) nuclease system. The latter, shown to have an interesting editing capacity, has become an essential tool for genome editing. In this review, we highlight the available literature on the methods and the applications of CRISPR-Cas for C. reinhardtii genetic engineering, including recent transformation methods, most used bioinformatic tools, best strategies for the expression of Cas protein and sgRNA, the CRISPR-Cas mediated gene knock-in/knock-out strategies, and finally the literature related to CRISPR expression and modification approaches

MOESM1 of Specific tracking of xylan using fluorescent-tagged carbohydrate-binding module 15 as molecular probe

Additional file 1: Figure S1. SDS-PAGE analysis of the OC15 probe purified by affinity chromatography. The expected molecular weight of the OC15 fusion protein is 44.68 kDa. A 12 % polyacrylamide gel was used for SDS-PAGE analysis. Well M: Precision plus protein standards (5 µg). Well OC15: Purified OC15 probe (10 µg)

Legislative Documents

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