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Comparing Heterotrophic with Phototrophic PHA Production
Published in Martin Koller, The Handbook of Polyhydroxyalkanoates, 2020
Ines Fritz, Katharina Meixner, Markus Neureiter, Bernhard Drosg
Currently achieved PHA concentrations are very low, so that strain improvement of cyanobacteria is intensively discussed. Gene amplification and gene transfer are the most promising techniques to achieve this goal. Unfortunately, cell densities in photo-autotrophic cultivation are lower by at least a factor of ten than in heterotrophic cultivation. With the same factor, the volumetric productivity is lower, and the investment costs are higher. Directly proportional to the reactor volume is the effort for downstream processing. Volumes of open ponds cannot be increased infinitely; the same is true for tube length or tube diameter in closed photobioreactors. Carbon dioxide saturation and O2 degassing is the limiting factor for all closed reactors. Reactor design needs to be optimized for energy efficiency, light use, flow management, mixing and distribution of cells and nutrients. The whole production plant needs to be designed for water and nutrient recycling and for biomass recovery, avoiding excessive need for finite resources. For example, coupling the photo-autotrophic growth with anaerobic digestion is a viable option. It seems to be obvious that acidic flue gas content, such as SO2 or NOX and toxic heavy metals, are unfavourable for photo-autotrophic microorganisms and need to be removed. Not so clear is the influence of solid particles (dust) or microbial contaminants (from biotechnological sources) on the performance and stability of a photo-autotrophic process.
Selection and Improvement of Industrial Organisms for Biotechnological Applications
Published in Nduka Okafor, Benedict C. Okeke, Modern Industrial Microbiology and Biotechnology, 2017
Nduka Okafor, Benedict C. Okeke
Two important features of plasmids to be used in genetic experiments may be compared by examining two plasmids. Plasmid psC101 has only two to five copies per cell and replicates with its host DNA. It is said to be under ‘stringent’ control. However, another plasmid pCol E 1 is found in about 25–30 copies per cell. It has a ‘relaxed control’ independent of the host and replicates without reference to the host DNA. When the host cell is starved of amino-acids or its protein synthesis is inhibited in some other manner, such as with the use of chloramphenicol, the Col E 1 plasmid continues to replicate for several hours until there are 1,000 to 3,000 copies per cell. Due to this high level of gene dosage (also referred to as gene amplification), products synthesized because of the presence of these plasmids are produced in extremely high amounts, a property of immense importance in biotechnology and industrial microbiology. Generally, conjugative plasmids are large, exhibit stringent control of DNA replication and are present in low copy numbers; on the other hand, non-conjugative plasmids are small, show relaxed DNA replication, and are present in high numbers (Table 7.5).
Vitamins and Nutrition
Published in Richard J. Sundberg, The Chemical Century, 2017
In the elderly, B12 deficiency can develop because of poor absorption by the digestive tract. In addition to anemia, there are also possible associations with cardiovascular disease, breast cancer, Alzheimer’s disease, and depression. Vitamin B12 for supplements is produced commercially using microorganisms. The microorganisms have been selected from mutated strains that are high producers. In recent years, further improvement in production has been achieved by gene amplification. Currently, the main producer is Aventis, using technology originally developed at Rhone-Polenc.4
A novel strategy to construct multi-strain starter cultures: an insight to evolve from natural to directed fermentation
Published in Preparative Biochemistry & Biotechnology, 2023
J. L. Navarrete-Bolaños, O. Serrato-Joya
Once the strain was selected, molecular identification was performed, which included ribosomal sequence analysis of the D1/D2 domains of the 28S rRNA gene and of the ITS region (ITS1: 5′-TCCGTAGGTGAACCTGCGG-3′) and ITS2: 5′-GCTGCGTTCTTCATCGATGC-3′, intergenic spaces and which includes the 5.8S rRNA gene). The method is based on genomic DNA extraction from selected strains, according to the protocol proposed by Ausubel et al.[30] using an Invitrogen™ column from the PureLink™ Genomic DNA set. For rRNA gene amplification, the extracted DNA was mixed with PCR Supermix (Invitrogen, Carlsbad, CA) containing high-fidelity DNA polymerase (Invitrogen). PCR was performed under the following conditions: initial denaturalization at 95 °C, 35 5-min cycles of 1 min of denaturation at 95 °C, 2 min of annealing at 50, 52, 54, 56, and 58 °C, and 2 min of extension at 72 °C, and a final extension for 10 min at 72 °C. The PCR products were purified with the QIAquick® Gel Extraction Kit (QIAGEN® Group). Once purified, the products were sent to the Laboratorio Nacional de Genómica para la Biodiversidad (LANGEBIO) of Cinvestav Irapuato, Gto. México for nucleotide sequencing, which was performed using the Sanger technique on an Applied Biosystems Automated 3730xl DNA Analyzer (Applied Biosystems by Life Technologies, Carlsbad, CA, USA) and a BigDye® Terminator v3.1 Cycle Sequencing Kit (Invitrogen). The sequence was compared with those reported in the National Center for Biotechnology Information (NCBI) database using the BLASTx algorithm for strain identification.
Tannin biodegradation by tannase from Serratia marcescens: optimization of production by response surface methodology and its partial characterization
Published in Chemical Engineering Communications, 2021
Amanda Reges de Sena, Lúzia Morgana de Melo Lopes, Miquéas Jamesse Gouveia, Marcos Juliano Gouveia, Marcelo Rodrigues Figueira de Mello, Tonny Cley Campos Leite, Gláucia Manoella de Souza Lima, Keila Aparecida Moreira, Sandra Aparecida de Assis
DNA extraction was performed by using culture grown in LB medium for 16 hours at 37 °C. After the growth period, the sample was centrifuged and DNA was extracted by using the Wizard Genomic DNA kit (Promega), according to the manufacturer’s instructions. The obtained DNA sequence was evaluated by agarose gel electrophoresis. Next, 16S rRNA gene amplification was performed by means of the DNA Polymerase Chain Reaction (PCR) technique using the following universal oligonucleotides (fD1 5′- AGAGTTTGATCCTGGCTCAG- 3′; rD1 5′- AAGGAGGTGATCCAGCC – 3′). It was used the reaction mixture consisted of 50 ng DNA, 10 pmol of each oligonucleotide, 200 mM of dNTP, 1.5 mM of MgCl2, 1X buffer, 1 U of Platinum Taq DNA polymerase (Invitrogen Life Technologies) in a final volume of 25 µL. The reaction condition was: 5 min denaturation at 94 °C, followed by 25 cycles of 1 min at 94 °C, 30 sec at 52 °C and 2 min at 72 °C, and a final extension of 10 min at 72 °C. The amplification product was analyzed by electrophoresis on agarose gel at 1.2% and the sample was subsequently sent to sequencing. The sample was sequenced by Macrogen and compared with all sequences in the GenBank by using Blast software from the National Center for Biotechnology Information (NCBI) (www.ncbi.nlm.nih.gov)
Overview of biological mechanisms of human carcinogens
Published in Journal of Toxicology and Environmental Health, Part B, 2019
Nicholas Birkett, Mustafa Al-Zoughool, Michael Bird, Robert A. Baan, Jan Zielinski, Daniel Krewski
Global hypo-methylation of DNA, combined with local hyper-methylation of selected genes, was noted in animal models and humans exposed to arsenic. Gene amplification was reported in some animal experiments. An adverse effect on DNA repair and p53 function was detected, although other studies reported increased DNA-repair activity, perhaps related to antioxidant effects in response to arsenic-induced oxidative damage. Arsenic stimulates an inflammatory response and interferes with apoptosis during chronic exposure. Arsenic was found to induce androgen independence in human prostate epithelial cells, which led to a 6-fold rise in expression of the K-RAS oncogene. Low levels of arsenic stimulate angiogenesis through an inflammatory process, leading to blood-vessel remodeling in the liver.