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Plasmid Sep 2019
Topics: Humans; Interspersed Repetitive Sequences; Plasmids
PubMed: 31472205
DOI: 10.1016/j.plasmid.2019.102437 -
Plasmid Mar 2005This review provides a historical perspective of the major findings that contributed to our current understanding of plasmid rolling-circle (RC) replication.... (Review)
Review
This review provides a historical perspective of the major findings that contributed to our current understanding of plasmid rolling-circle (RC) replication. Rolling-circle-replicating (RCR) plasmids were discovered approximately 20 years ago. The first of the RCR plasmids to be identified were native to Gram-positive bacteria, but later such plasmids were also identified in Gram-negative bacteria and in archaea. Further studies revealed mechanistic similarities in the replication of RCR plasmids and the single-stranded DNA bacteriophages of Escherichia coli, although there were important differences as well. Three important elements, a gene encoding the initiator protein, the double strand origin, and the single strand origin, are contained in all RCR plasmids. The initiator proteins typically contain a domain involved in their sequence-specific binding to the double strand origin and a domain that nicks within the double strand origin and generates the primer for DNA replication. The double strand origins include the start-site of leading strand synthesis and contain sequences that are bound and nicked by the initiator proteins. The single strand origins are required for synthesis of the lagging strand of RCR plasmids. The single strand origins are non-coding regions that are strand-specific, and contain extensive secondary structures. This minireview will highlight the major findings in the study of plasmid RC replication over the past twenty years. Regulation of replication of RCR plasmids will not be included since it is the subject of another review.
Topics: Base Sequence; DNA; DNA Replication; DNA, Single-Stranded; Forecasting; Molecular Sequence Data; Plasmids
PubMed: 15737400
DOI: 10.1016/j.plasmid.2004.12.008 -
Recent Patents on Biotechnology 2008The demand for plasmid DNA has increased vastly in response to rapid advances in its use in gene therapy and vaccines. These therapies are based on the same principle,... (Review)
Review
The demand for plasmid DNA has increased vastly in response to rapid advances in its use in gene therapy and vaccines. These therapies are based on the same principle, i.e. the introduction of nucleic acids in human/non-human cells receptor to restore, cancel, enhance or introduce a biochemical function. Naked plasmid DNA as a vector has attracted a lot of interest since it offers several advantages over a viral vector, especially weak immunogenicity, better safety and easy to manufacture, but low transfection efficacy. Non-viral gene therapy may require considerable amounts (milligram scale) of pharmaceutical-grade pDNA per patient since the efficacy and duration of gene expression is presently relatively low. Reliance on fermentation, which generates large lysate volumes, for producing the needed quantities of pDNA is becoming more widespread. Through optimization of the biological system, growth environment and the growth mode, improvements can be achieved in biomass productivity, plasmid yield, plasmid quality and production costs. The information on large-scale plasmid production is scarce and usually not available to the scientific community. This review summarizes recent patents and patent applications relating to plasmid upstream processing manufacturing, ranging from plasmid design to growth strategies to produce plasmid-bearing E. coli.
Topics: Drug Industry; Genetic Therapy; Patents as Topic; Plasmids; Vaccines, DNA
PubMed: 19075863
DOI: 10.2174/187220808786241015 -
Methods in Molecular Medicine 2006A large-scale approach to the purification of plasmid DNA has been developed that overcomes many of the limitations of current chromatography-based processes. The... (Review)
Review
A large-scale approach to the purification of plasmid DNA has been developed that overcomes many of the limitations of current chromatography-based processes. The process consists of a scaleable lysis using recombinant lysozyme and a rapid heating and cooling step followed by a selective precipitation with cetyltrimethylammonium bromide (CTAB). Calcium silicate batch adsorption is then utilized to remove residual genomic DNA, linear plasmid, open circular plasmid, endotoxin, detergents, and proteins. Finally, a concentration and diafiltration step utilizing ultrafiltration and a terminal sterile filtration complete the process. The final product exceeds the requirements for clinical-grade plasmid DNA, and the process has been scaled up to yield an average of 18 +/- 4 g (over five lots) of pharmaceutically pure plasmid DNA per 140 L of lysate (from approx 1.3 kg Escherichia coli dry cell weight).
Topics: Calcium Compounds; Cetrimonium; Cetrimonium Compounds; Escherichia coli; Muramidase; Plasmids; Silicates
PubMed: 16988465
DOI: 10.1385/1-59745-168-1:351 -
Current Opinion in Molecular... Aug 2003Over the past several years, significant progress has been made in the development of non-viral methodologies that can effectively deliver genes to target tissues in... (Review)
Review
Over the past several years, significant progress has been made in the development of non-viral methodologies that can effectively deliver genes to target tissues in vivo. One of the most surprising successes has been the discovery that naked plasmid DNA (pDNA) can be delivered into tissues such as liver and muscle with high efficiency using the vascular system. The key breakthrough involved the realization that pDNA could be injected rapidly into blood vessels (using increased volumes) in a manner that facilitates extravasation of the DNA solution outside the blood vessel wall. The extravasation process places the DNA in contact with the plasma membranes of the underlying parenchymal cells of the target organ. This intravascular delivery technique, termed 'hydrodynamic delivery', has become established as the primary non-viral methodology for delivering pDNA expression constructs to target tissues in vivo. This review highlights many of the most recent studies in which increased volume/rapid injection procedures have been used. These include studies in which the technology was used as a new and powerful tool to address in vivo gene expression questions, as well as numerous studies that were designed to better understand or improve the methodology. It is these scientific efforts that have served to fuel the development of this delivery technology from simply an interesting phenomenon to a highly useful and broadly used gene delivery methodology.
Topics: Animals; Drug Delivery Systems; Genetic Therapy; Humans; Injections, Intravenous; Plasmids
PubMed: 14513675
DOI: No ID Found -
Journal of Pharmaceutical Sciences Jul 2002The majority of gene therapy clinical trials use plasmid DNA that is susceptible to shear-induced degradation. Many processing steps in the extraction, purification, and... (Review)
Review
The majority of gene therapy clinical trials use plasmid DNA that is susceptible to shear-induced degradation. Many processing steps in the extraction, purification, and preparation of plasmid-based therapeutics can impart significant shear stress that can fracture the phosphodiester backbone of polynucleotides, and reduce biological activity. Much of the mechanistic work on shear degradation of DNA was conducted over 30 years ago, and we rely heavily on this early work in an attempt to explain the empirical observations of more recent investigations concerning the aerosolization of plasmids. Unfortunately, the sporadic reports of shear degradation in the literature use different experimental systems, making it difficult to quantitatively compare results and reach definitive mechanistic conclusions. In this review, we describe the forces imparted to DNA during shear stress, and use published data to quantitatively evaluate their relative effects. In addition, we discuss the effects of molecular weight, strain rate, particle size, flexibility, ionic strength, gas-liquid interfaces, and turbulence on the fluid flow degradation of supercoiled plasmid DNA. Finally, we speculate on computational methods that might allow degradation rates in different experimental systems to be predicted.
Topics: Animals; Biomechanical Phenomena; DNA, Circular; Genetic Therapy; Humans; Plasmids; Shear Strength
PubMed: 12115820
DOI: 10.1002/jps.10140 -
Infection and Immunity Jul 2011Virulence functions of pathogenic bacteria are often encoded on large extrachromosomal plasmids. These plasmids are maintained at low copy number to reduce the metabolic... (Review)
Review
Virulence functions of pathogenic bacteria are often encoded on large extrachromosomal plasmids. These plasmids are maintained at low copy number to reduce the metabolic burden on their host. Low-copy-number plasmids risk loss during cell division. This is countered by plasmid-encoded systems that ensure that each cell receives at least one plasmid copy. Plasmid replication and recombination can produce plasmid multimers that hinder plasmid segregation. These are removed by multimer resolution systems. Equitable distribution of the resulting monomers to daughter cells is ensured by plasmid partition systems that actively segregate plasmid copies to daughter cells in a process akin to mitosis in higher organisms. Any plasmid-free cells that still arise due to occasional failures of replication, multimer resolution, or partition are eliminated by plasmid-encoded postsegregational killing systems. Here we argue that all of these three systems are essential for the stable maintenance of large low-copy-number plasmids. Thus, they should be found on all large virulence plasmids. Where available, well-annotated sequences of virulence plasmids confirm this. Indeed, virulence plasmids often appear to contain more than one example conforming to each of the three system classes. Since these systems are essential for virulence, they can be regarded as ubiquitous virulence factors. As such, they should be informative in the search for new antibacterial agents and drug targets.
Topics: Bacteria; Bacterial Proteins; Cell Division; Conjugation, Genetic; DNA Replication; DNA, Bacterial; Gene Dosage; Plasmids; Virulence Factors
PubMed: 21555398
DOI: 10.1128/IAI.00127-11 -
Methods in Molecular Medicine 2006Qiagen offers a unique technology for plasmid manufacturing, working reliably for every parent plasmid. The process steps such as strain and clone selection, and... (Review)
Review
Qiagen offers a unique technology for plasmid manufacturing, working reliably for every parent plasmid. The process steps such as strain and clone selection, and fermentation optimization ensure optimal plasmid DNA yield and quality in the starting material. Master Cell Bank and Working Cell Bank manufacturing is then performed under cGMP conditions. A high-yield, low mechanical stress alkaline lysis procedure, followed by a proprietary endotoxin-removal step and anion-exchange chromatography ensures consistently high plasmid DNA quality. The material undergoes stringent quality control tests and is accompanied by a comprehensive quality control report and a documentation package for regulatory filing. The following chapter describes the necessary steps such as host cell selection, growth conditions, downstream processing, and quality assurance and control.
Topics: Chromatography, Ion Exchange; Escherichia coli K12; Guidelines as Topic; Industrial Microbiology; Plasmids; Quality Control
PubMed: 16988464
DOI: 10.1385/1-59745-168-1:339 -
Journal of Pharmaceutical Sciences Feb 1998The advent of gene therapy and polynucleotide-based vaccines has resulted in the use of plasmid DNA as a drug substance. Although biologically (cell or animal) based... (Comparative Study)
Comparative Study Review
The advent of gene therapy and polynucleotide-based vaccines has resulted in the use of plasmid DNA as a drug substance. Although biologically (cell or animal) based assays must currently be employed to establish the identity and potency of such drugs, we argue that in the future, a combination of microchip-based mutation detection devices combined with an array of chromatographic, electrophoretic, hydrodynamic, and spectroscopic methods can be employed to rigorously establish these properties. We review a variety of such methods in this context and also consider the issue of the chemical stability of plasmids. Extensive comparison is made to protein-based pharmaceuticals with the unique importance of polynucleotide sequence emphasized in comparison to protein tertiary structure.
Topics: Chemistry Techniques, Analytical; Chemistry, Pharmaceutical; DNA Mutational Analysis; Drug Stability; Freeze Etching; Genetic Techniques; Light; Microscopy, Atomic Force; Microscopy, Electron; Molecular Structure; Nucleic Acid Denaturation; Particle Size; Plasmids; Protein Structure, Tertiary; Scattering, Radiation
PubMed: 9519144
DOI: 10.1021/js970367a -
Nature Reviews. Microbiology Jan 2022
Topics: Plasmids
PubMed: 34697498
DOI: 10.1038/s41579-021-00657-x