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Nanopharmaceuticals in Cardiovascular Medicine
Published in Harishkumar Madhyastha, Durgesh Nandini Chauhan, Nanopharmaceuticals in Regenerative Medicine, 2022
Ramandeep Singh, Anupam Mittal, Maryada Sharma, Ajay Bahl, Madhu Khullar
Besides being formed with the naturally present bilipid layer, they might not be able to penetrate deep into the skin when administered through the transdermal route. This can be enhanced by the incorporation of some anionic surfactants and ethanol which can increase the fluidity of the bilipid layer, thus enhancing the penetration of the liposomes. Earlier for gene transfer, DNA or gene of interest was assembled with liposomes but was prone to degradation by endocytosis mediated transfer. Thus, cationic liposomes were developed. Electrostatic interactions between positive cationic liposomes and DNA, which are negatively charged, stabilize the complex and improve the rate of transfection as well. These positively charged cationic liposomes also protected the DNA or gene of interest against degradation (Marcato et al., 2008).
Basic Microbiology
Published in Philip A. Geis, Cosmetic Microbiology, 2020
Unlike eukaryotic cells, bacteria can transfer DNA sequences between individuals that are not of the same lineage. This can occur between members of the same species or between different species. The three main mechanisms that bacteria use to perform horizontal gene transfer are listed here. Transformation—DNA found within the extracellular environment of the bacteria is actively taken up into the bacteria and is used either in its native state, such as with plasmids, or incorporated into the genome, as with chromosomal DNA sequences. The utilization of these sequences can often confer drastic new phenotypes on the bacteria that harbor those sequences.Transduction—Viruses that are present, incorporated into the chromosome of a bacterium, exit the infected bacteria to infect new host cells. In the process, they remove small portions of the chromosome of the infected bacteria. In the new host, these sequences are integrated into the new host genome.Conjugation—As mentioned previously, bacteria that are able to conjugate by producing sex pili can transfer DNA sequences across the pili and lead to incorporation of new chromosomal sequences within the other mating bacteria.
Methods of Evaluation in Orthopaedic Animal Research
Published in Yuehuei H. An, Richard J. Friedman, Animal Models in Orthopaedic Research, 2020
Hybridization is the process of matching complementary strands of DNA or RNA or both to form a double stranded molecule. In situ hybridization (ISH) is the hybridization of a DNA or RNA probe to a target molecule that has not been extracted from its original cellular location, within a chromosome or in a fixed tissue section.253 Immunohistochemistry is analogous to ISH for nucleic acids and is used to detect the distribution of a specific protein within a cell or tissue. In immunohistochemistry, a specific antibody serves as the probe to detect the protein of interest. Gel electrophoresis is a method of separating DNA, RNA, or protein molecules based on their size and electrical charge. This technique makes use of the fact that, under an electrical field small molecules migrate through a gel matrix (agarose or acrylamide) faster than larger molecules. Southern blotting or transfer is a technique which is used to transfer DNA that has been electrophoresed through an agarose gel onto a solid support for hybridization. Northern blotting or transfer is the process of transferring RNA onto a solid filter support for hybridization. Western blotting or transfer is the process of transferring proteins that have been electrophoresed through an acrylamide gel onto a solid filter support for detection of a specific protein by antibody labeling.
How to discover new antibiotic resistance genes?
Published in Expert Review of Molecular Diagnostics, 2019
Linda Hadjadj, Sophie Alexandra Baron, Seydina M. Diene, Jean-Marc Rolain
The highlighting of an ARG transfer by conjugation is a reference method and is still used today to identify new ARGs [17–19]. A donor strain resistant to a specific antibiotic can confer resistance to a receptor strain initially susceptible to this antibiotic, and resistant mutant are selected by using this antibiotic as selection marker. This AR transfer is possible via a plasmid carrying an ARG. This natural mechanism is the main mechanism responsible for the spread of antibiotic resistance [20]. Plasmid transfer can also be performed by transformation, if the plasmid is not conjugative. Transduction is a third method used to transfer bacterial DNA from a donor strain to a receptor strain via a bacteriophage. These mechanisms are currently used by bacteria to transfer DNA to other bacteria and have been reproduced in vitro to highlight the presence of a transferrable gene.
Two unresolved issues in community engagement for field trials of genetically modified mosquitoes
Published in Pathogens and Global Health, 2019
Both methods have limitations and potential environmental and public health risks. A risk that both methods have in common is that the GM mosquitoes might migrate out of the release site [1–3]. However, this risk can be minimized with proper site selection and containment measures [1–3]. The most significant risk of the method that suppresses the targeted mosquito population is that it could have adverse effects on the food web, because some species of bats, birds, amphibians, insects, and fish eat mosquitoes in adult or larval forms. However, it is likely that impacted predator species would find substitute food sources, so the food web would adjust [1–3,13,20]. Another potential adverse outcome that the lethal gene could have harmful impacts on non-target species if it is transferred to non-target species by horizontal gene transfer and is expressed.3Horizontal gene transfer is a process in which DNA moves between organisms other than by sexual reproduction. Viruses can transfer DNA to numerous species, including humans. Horizontal gene transfer can transfer DNA across species [3]. However, the FDA has determined that the probability of this type of adverse outcome is very low because horizontal gene transfer is a rare event [20]. While some citizens and health-care professionals have been concerned that the GM mosquitoes could transfer the lethal gene to human beings, this risk is negligible because male mosquitoes do not bite [20]. The main limitation of this method is that it would need to be re-implemented periodically to keep targeted mosquito populations in check [3,13,20].
Application in Gene Editing in Ovarian Cancer Therapy
Published in Cancer Investigation, 2022
Shuang Luo, Yujiao Wang, Yongyu Tao, Shuo Li, Zirui Wang, Wei He, Hangxing Wang, Nan Wang, Jianwei Xu, Hailiang Song
Gene editing is a new technique for the site-specific modification of the genome, which can be used to accurately locate a specific site in the genome, where a target DNA fragment can be cut and a new gene fragment can be inserted. This process not only stimulates the natural mutation of genes but also modifies the genome in a specific manner. The conventional method to obtain gene mutants is to construct a large-scale random mutant library by using transfer DNA or transposons, but the construction of saturated mutants that cover the whole genome is time consuming and requires a high workload (10,11). In recent years, with the emergence of highly specific and operable artificial nucleases, gene editing has rapidly improved. At present, target genes can be completely inactivated by site-directed mutation, and then a series of operations can be carried out. Compared to the traditional method of acquiring mutants, this is the most direct and effective method to study the function of a specific gene. Modern gene editing technologies share the same basic principle, i.e., activation of the natural repair mechanism of the cell through specific DNA double-stranded breaks (DSBs), which mainly includes two pathways: non-homologous end joining (NHEJ) and homologous recombinant repair (12). NHEJ has a high probability of mismatch (13). In the repair and reconnection process of DSBs, random insertion or loss of bases can occur, which results in frameshift mutations due to nucleases and gene inactivation, indirectly achieving knockout of the target gene. If an exogenous donor gene sequence is present, the NHEJ mechanism will connect it with the DSB site, thereby enabling targeted knock in. Homologous recombinant repair is a relatively high-fidelity repair process. In the presence of a predominant donor with a homologous arm, the exogenous target genes in the donor will be completely integrated into the target site through the homologous recombination process without random base insertion or loss (10). If the DSB is produced on both sides of a gene, gene replacement can be performed in the presence of a homologous donor. According to the different nucleases used, gene editing can be divided into three types: zinc-finger nucleases (ZFNs) mediated by artificial nucleic acids, transcription activator-like effector nuclease (TALEN) technology, and clustered regularly interspaced short palindromic repeats (CRISPR) and their associated protease 9 (Cas9) RNA-guided nuclease (RGN) technology.