Genetic manipulation of microalgae for enhanced biotechnological applications

2022 ◽  
pp. 97-122
Author(s):  
Ashutosh Pandey ◽  
Gaurav Kant ◽  
Shadma Afzal ◽  
Manish Pratap Singh ◽  
Nand Kumar Singh ◽  
...  
Keyword(s):  
Genetic Manipulation ◽  
Biotechnological Applications

scholarly journals Yeast Surface Display System: Strategies for Improvement and Biotechnological Applications

2022 ◽  
Vol 9 ◽  
Author(s):  
Karla V. Teymennet-Ramírez ◽  
Fernando Martínez-Morales ◽  
María R. Trejo-Hernández
Keyword(s):  
Protein Interactions ◽  
Genetic Manipulation ◽  
Surface Display ◽  
Yeast Surface Display ◽  
Yeast Cells ◽  
Biotechnological Applications ◽  
Protein Protein Interactions ◽  
Post Translational Modifications ◽  
Yeast Surface ◽  
Antibody Design

Yeast surface display (YSD) is a “whole-cell” platform used for the heterologous expression of proteins immobilized on the yeast’s cell surface. YSD combines the advantages eukaryotic systems offer such as post-translational modifications, correct folding and glycosylation of proteins, with ease of cell culturing and genetic manipulation, and allows of protein immobilization and recovery. Additionally, proteins displayed on the surface of yeast cells may show enhanced stability against changes in temperature, pH, organic solvents, and proteases. This platform has been used to study protein-protein interactions, antibody design and protein engineering. Other applications for YSD include library screening, whole-proteome studies, bioremediation, vaccine and antibiotics development, production of biosensors, ethanol production and biocatalysis. YSD is a promising technology that is not yet optimized for biotechnological applications. This mini review is focused on recent strategies to improve the efficiency and selection of displayed proteins. YSD is presented as a cutting-edge technology for the vectorial expression of proteins and peptides. Finally, recent biotechnological applications are summarized. The different approaches described herein could allow for a better strategy cascade for increasing protein/peptide interaction and production.

scholarly journals Incorporation of Magnetic Nanoparticles into Protoplasts of Microalgae Haematococcus pluvialis: A Tool for Biotechnological Applications

Molecules ◽  
2020 ◽  
Vol 25 (21) ◽  
pp. 5068
Author(s):  
Maria G. Savvidou ◽  
Angelo Ferraro ◽  
Evangelos Hristoforou ◽  
Diomi Mamma ◽  
Dimitris Kekos ◽  
...  
Keyword(s):  
Cell Wall ◽  
Magnetic Nanoparticles ◽  
Mechanical Treatment ◽  
Genetic Manipulation ◽  
Haematococcus Pluvialis ◽  
Biotechnological Applications ◽  
Protoplast Formation ◽  
Cell Recovery ◽  
Microalgae Biomass ◽  
A Cell

Intensive research on the use of magnetic nanoparticles for biotechnological applications of microalgae biomass guided the development of proper treatment to successfully incorporate them into these single-cell microorganisms. Protoplasts, as cells lacking a cell wall, are extensively used in plant/microalgae genetic manipulation as well as various biotechnological applications. In this work, a detailed study on the formation of protoplasts from Haematococcus pluvialis with the use of enzymatic and mechanical procedures was performed. The optimization of several parameters affecting the formation of protoplasmic cells and cell recovery was investigated. In the enzymatic treatment, a solution of cellulase was studied at different time points of incubation, whereas in the mechanical treatment, glass beads vortexing was used. Mechanical treatment gave better results in comparison to the enzymatic one. Concerning the cell recovery, after the protoplast formation, it was found to be similar in both methods used; cell viability was not investigated. To enhance the protoplast cell wall reconstruction, different “recovery media” with an organic source of carbon or nitrogen were used. Cell morphology during all treatments was evaluated by electron microscopy. The optimal conditions found for protoplast formation and cell reconstruction were successfully used to produce Haematococcus pluvialis cells with magnetic properties.

scholarly journals Marker-free genome engineering in Amycolatopsis using the pSAM2 site-specific recombination system

2021 ◽  
Author(s):  
Luísa D. F. Santos ◽  
Laëtitia Caraty-Philippe ◽  
Emmanuelle Darbon ◽  
Jean-Luc Pernodet
Keyword(s):  
Genome Engineering ◽  
High Efficiency ◽  
Genetic Manipulation ◽  
Marker Genes ◽  
Biotechnological Applications ◽  
Site Specific ◽  
Site Specific Recombination ◽  
Recombination System ◽  
Marker Free ◽  
Site Specific Recombination System

ABSTRACTActinobacteria belonging to the genus Amycolatopsis are important for antibiotic production and other valuable biotechnological applications such as biodegradation or bioconversion. Despite their industrial importance, tools and methods for the genetic manipulation of Amycolatopsis are less developed than in other actinobacteria such as Streptomyces. Moreover, most of the existing methods do not support convenient marker-free genome engineering. Here, we report the use of the pSAM2 site-specific recombination system for the efficient deletion of marker genes or large DNA regions in Amycolatopsis. For this purpose, we constructed a shuttle vector, replicating in Escherichia coli and Amycolatopsis, expressing the Xis and Int proteins from the Streptomyces integrative and conjugative element pSAM2. These proteins are sufficient for site-specific recombination between the attachment sites attL and attR. We also constructed two plasmids, replicative in E. coli but not in Amycolatopsis, for the integration of the recombination sites attL and attR on each side of a region targeted for deletion. We exemplified the use of these tools in Amycolatopsis mediterranei DSM 40773 by obtaining with high efficiency (>95%) a marker-free deletion of one single gene in the rifamycin biosynthetic gene cluster or of the entire 90-kb cluster.IMPORTANCEThe genus Amycolatopsis is regarded as an important source of diverse specialized metabolites. Members of this genus are used in industry for the production of valuable antibiotics such as rifamycins or vancomycin. Amycolatopsis spp. also present a great interest for biotechnological applications such as biodegradation or bioconversion. Despite their importance, their genetic manipulation was somehow hampered by the lack of efficient tools. Here we report the successful use of the pSAM2 site-specific recombination system to construct unmarked deletion mutants, allowing marker recycling, or to create large deletions in A. mediterranei DSM 40773. The high efficiency of this site-specific recombination system and it possible application to other Amycolatopsis species open new opportunities for marker-free genome engineering in this genus.

Metabolic engineering of CHO cells to prepare glycoproteins

2018 ◽  
Vol 2 (3) ◽  
pp. 433-442 ◽  
Author(s):  
Qiong Wang ◽  
Michael J. Betenbaugh
Keyword(s):  
Metabolic Engineering ◽  
Cho Cells ◽  
Genetic Manipulation ◽  
Chinese Hamster ◽  
Post Translational Modification ◽  
Effector Functions ◽  
Recombinant Glycoproteins ◽  
Production Platform ◽  
Chemical Inhibitors ◽  
Amino Acid Mutations

As a complex and common post-translational modification, N-linked glycosylation affects a recombinant glycoprotein's biological activity and efficacy. For example, the α1,6-fucosylation significantly affects antibody-dependent cellular cytotoxicity and α2,6-sialylation is critical for antibody anti-inflammatory activity. Terminal sialylation is important for a glycoprotein's circulatory half-life. Chinese hamster ovary (CHO) cells are currently the predominant recombinant protein production platform, and, in this review, the characteristics of CHO glycosylation are summarized. Moreover, recent and current metabolic engineering strategies for tailoring glycoprotein fucosylation and sialylation in CHO cells, intensely investigated in the past decades, are described. One approach for reducing α1,6-fucosylation is through inhibiting fucosyltransferase (FUT8) expression by knockdown and knockout methods. Another approach to modulate fucosylation is through inhibition of multiple genes in the fucosylation biosynthesis pathway or through chemical inhibitors. To modulate antibody sialylation of the fragment crystallizable region, expressions of sialyltransferase and galactotransferase individually or together with amino acid mutations can affect antibody glycoforms and further influence antibody effector functions. The inhibition of sialidase expression and chemical supplementations are also effective and complementary approaches to improve the sialylation levels on recombinant glycoproteins. The engineering of CHO cells or protein sequence to control glycoforms to produce more homogenous glycans is an emerging topic. For modulating the glycosylation metabolic pathways, the interplay of multiple glyco-gene knockouts and knockins and the combination of multiple approaches, including genetic manipulation, protein engineering and chemical supplementation, are detailed in order to achieve specific glycan profiles on recombinant glycoproteins for superior biological function and effectiveness.

Drug-Induced Agranulocytosis: A Mini Review

2016 ◽  
Vol 2 (1) ◽  
pp. 57-59
Author(s):  
Pavithra D ◽  
Praveen D ◽  
Vijey Aanandhi M
Keyword(s):  
Bone Marrow ◽  
Complete Blood Count ◽  
Genetic Manipulation ◽  
White Blood Cells ◽  
Deep Infection ◽  
Morbidity Rate ◽  
Drug Induced ◽  
Serious Adverse Event ◽  
Mortality And Morbidity ◽  
The Individual

Agranulocytosis is also known to be granulopenia, causing neutropenia in circulating blood streams .The destruction of white blood cells takes place which leads to increase in the infection rate in an individual where immune system of the individual is suppressed. The symptoms includes fever, sore throat, mouth ulcers. These are commonly seen as adverse effects of a particular drug and are prescribed for the common diagnostic test for regular monitoring of complete blood count in an admitted patient. Drug-induced agranulocytosis remains a serious adverse event due to occurrence of severe sepsis with deep infection leading to pneumonia, septicaemia, and septic shock in two/third of the patient. Antibiotics seem to be the major causative weapon for this disorder. Certain drugs mainly anti-thyroid drugs, ticlopidine hydrochloride, spironolactone, clozapine, antileptic drugs (clozapine), non-steroidal anti-inflammatory agents, dipyrone are the potential causes. Bone marrow insufficiency followed by destruction or limited proliferative bone marrow destruction takes place. Chemotherapy is rarely seen as a causative agent for this disorder. Genetic manipulation may also include as one of the reason. Agranulocytosis can be recovered within two weeks but the mortality and morbidity rate during the acute phase seems to be high, appropriate adjuvant treatment with broad-spectrum antibiotics are prerequisites for the management of complicated neutropenia. Drugs that are treated for this are expected to change as a resistant drug to the patient. The pathogenesis of agranulocytosis is not yet known. A comprehensive literature search has been carried out in PubMed, Google Scholar and articles pertaining to drug-induced agranulocytosis were selected for review.

Improvement of effectiveness in maize breeding

2006 ◽  
Vol 54 (3) ◽  
pp. 351-358 ◽  
Author(s):  
P. Pepó
Keyword(s):  
Recurrent Selection ◽  
Genetic Manipulation ◽  
Field Testing ◽  
Tissue Cultures ◽  
Maize Breeding ◽  
Breeding Programmes ◽  
Speed Up ◽  
Selection For

Plant regeneration via tissue culture is becoming increasingly more common in monocots such as maize (Zea mays L.). Pollen (gametophytic) selection for resistance to aflatoxin in maize can greatly facilitate recurrent selection and the screening of germplasm for resistance at much less cost and in a shorter time than field testing. In vivo and in vitro techniques have been integrated in maize breeding programmes to obtain desirable agronomic attributes, enhance the genes responsible for them and speed up the breeding process. The efficiency of anther and tissue cultures in maize and wheat has reached the stage where they can be used in breeding programmes to some extent and many new cultivars produced by genetic manipulation have now reached the market.

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