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Introduction to Cells, DNA, and Viruses
Published in Patricia G. Melloy, Viruses and Society, 2023
Experiments done by scientists in the 1940s and 1950s determined that the DNA is the hereditary material in structures within the nucleus called chromosomes. Chromosomes also are made of protein, which plays a supporting structural role in the chromosomes. The DNA contains the hereditary units known as genes. The researchers Oswald Avery, Colin MacLeod, and Maclyn McCarty performed a series of experiments, an extension of experiments done by Frederick Griffith, looking at a phenomenon called bacterial transformation. Transformation in this context is the transfer of genes from one bacterium into another, with the bacterium now being transformed or changed in its heritable characteristics. They determined that DNA was responsible for the transformation (Avery, MacLeod, and McCarty 1944). Interestingly, scientists sometimes use harmless viruses as a tool in their research. In the famously elegant Hershey and Chase experiment done in 1952, researchers used a type of virus known to infect bacteria called a bacteriophage. A radioactive phosphorus label was used to track the DNA from the virus, and a radioactive sulfur label was used to track the protein of the virus. (There is no sulfur in DNA and no phosphorus in protein, so the labels are specific to those macromolecules.) These researchers discovered that the DNA was taken into the infected cells, and it encoded the genetic material needed to make copies of the bacteriophage in the host cells (Hershey and Chase 1952).
Gene Therapy in Oral Tissue Regeneration
Published in Vincenzo Guarino, Marco Antonio Alvarez-Pérez, Current Advances in Oral and Craniofacial Tissue Engineering, 2020
Fernando Suaste, Patricia González-Alva, Alejandro Luis, Osmar Alejandro
In bacteria, the delivery of genetic material has been done through the use of plasmids, which are circular DNA molecules that replicate and segregate independently on the host bacterial chromosome (episomes); this allows that the sequences cloned in these vectors are expressed stablely in the bacterial cytosol. Bacterial transformation emerged as one of the first approaches to introduce genetic material into a cell. In this strategy the permeability of the cell membrane is disturbed either by thermal shock or by the use of an electric field (electroporation). Additionally, cellular mechanisms such as conjugation and phage-mediated transduction were used to transfer genetic elements from one cell to another.
Biology of microbes
Published in Philip A. Geis, Cosmetic Microbiology, 2006
The most common way bacteria accomplish this task is via general recombination involving a reciprocal exchange between pairs of homologous DNA sequences. It occurs anywhere on the chromosome where a homology exists with another DNA strand where a break and subsequent cross-over can occur. Many of the enzymes involved in carrying out this recombination are the same as those involved in DNA repair. In bacterial transformation (detailed below), a nonreciprocal recombination occurs where sections of genetic material are inserted into the chromosome by incorporating a single strand of DNA that forms a stretch of heteroduplex DNA. Although the one strand is homologous, it is inserted into the genome as a complementary strand to the original and not duplicated.
Tracing the origins of extracellular DNA in bacterial biofilms: story of death and predation to community benefit
Published in Biofouling, 2021
Davide Campoccia, Lucio Montanaro, Carla Renata Arciola
In Neisseria gonorrhoeae, secretion of chromosomal DNA has been found to be associated to a type IV secretion system (TFSS) implicated in bacterial transformation. Salgado-Pabón et al. (2010) observed that, within the cell population, gonococcal variants that produce type IV pili release larger quantities of DNA than non-piliated variants. Piliated strains would produce the largest quantity of DNA secretion in late log-phase growth and DNA release would not occur through autolysis. Moreover, secreted DNA would be in the form of single-strand DNA protected at the 5′ end from 5′–3′ exonuclease digestion (Salgado-Pabón et al. 2007). Secretion would occur between the 2-h and 2.5-h time points and precede the onset of the stationary/death phase during which bacteria would switch to cell lytic processes.
Engineering of siRNA loaded PLGA Nano-Particles for highly efficient silencing of GPR87 gene as a target for pancreatic cancer treatment
Published in Pharmaceutical Development and Technology, 2020
Seyma Ceylan, Fatemeh Bahadori, Fahri Akbas
RPMI-1640 medium (Gibco®, Invitrogen, Loughborough- UK), supplemented with 10% inactivated fetal bovine serum (FBS), was purchased from Gibco-BRL (Cheshire, UK), penicillin (100 U/mL) and streptomycin (100 mg/mL) from Invitrogen (Renfrew, UK). Rapid DNA Ligation Kit, TransformAid Bacterial Transformation Kit, and PureLink™ HiPure Plasmid Miniprep Kit were obtained from Thermo Scientific (Renfrew, UK). Dual-Luciferase® Reporter Assay System and FuGENE HD Transfection Reagent was obtained from Promega (Madison, USA), HEK293T cells originally referred as 293tsA1609neo, is a highly transfectable derivative of human embryonic kidney 293 cells were purchased from ATCC® (CRL-3216) (VA-USA) and 1.1 B4 Cell, is human pancreatic Beta cell: PANC-1 hybrid cell line which is insulin-secreting cell line were purchased from Sigma-Aldrich (Dorset, UK) 10012801.
The potential protective roles of zinc, selenium and glutathione on hypoxia-induced TRPM2 channel activation in transfected HEK293 cells
Published in Journal of Receptors and Signal Transduction, 2020
Dilek Duzgun Ergun, Sefik Dursun, Nural Pastaci Ozsobaci, Ozden Hatırnaz Ng, Mustafa Naziroglu, Dervis Ozcelik
Pre-prepared E. coli DH5α competent cells were used for bacterial transformation of pcDNA3-IRES-EGFP-TRPM2 plasmid (TRPM2 gene). The plasmids were transformed into competent cells through heat shock. In order to determine the right colony that contains the desired plasmid, 6 different colonies were selected from the petri dishes where reproduction was observed. The plasmids were isolated from each colony using Miniprep Kit (Qiagen, Germany), and following the kit protocol. Quality control was performed by running the isolated plasmids on 1% agarose gel electrophoresis.