Of plasmids

Hansma H G, Vesenka J, Siegerist C, Kelderman G, Morrett H, Sinsheimer R L, Bustamante C, Elings V and Hansma P K 1992 Reproducible imaging and dissection of plasmid DNA under liquid with the atomic force microscope Science 256 1180... 

Most plasmids are topologically closed circles of DNA. They can be separated from the bulk of the chromosomal DNA by virtue of their resistance to alkaline solution. The double-stranded stmcture of DNA is denatured at high pH, but because the two strands of the plasmid are topologically joined they are more readily renatured. This property is exploited in rapid procedures for the isolation of plasmid DNA from recombinant microorganisms (5,6). 

Plasmid Vectors for Facile Introduction of Passenger DNA and Selection of Recombinants. The map of a commonly used plasmid vector, pUC19 (7), is shown in Figure 2. Three parts of the vector are key to its utility. The origin sequence, oh, allows the repHcation of plasmid DNA in high copy number relative to the chromosome. A gene, amp, encoding the enzyme beta-lactamase, which hydrolyzes penicillin compounds, allows... 

Toyopearl HW-75 resin, with pores larger than 1000 A, have been used in place of ultracentrifugation steps for the purification of plasmid DNA. Ultracentrifugation is a time-consuming process and requires expensive chemicals, such as cesium chloride. Toyopearl HW-75 resin provides superior separation performance for plasmid DNA and also provides high yields (54). 

Many copies of plasmid widi desired site-specific... 

Continuous. Long fermentation run times, with many generations, increase the chance of mutation or loss of plasmid DNA. 

Liu MA, Ulmer JB (2005) Human clinical trials of plasmid DNA vaccines. Adv Genet 55 25-40... 

Resistance to trimethoprim can be due to the acquisition of plasmid encoded non-allelic variants of the chromosomal DHFR enzyme that are antibiotic unsusceptible. The genes may be part of transposons that then insert into the chromosome. For instance, in gram-negative bacteria the most widespread gene is dhfrl on transposon Tn7. 

The transfection mechanism of plasmid-chitosan complexes as well as the relationship between transfection activity and cell uptake was analyzed by using fluorescein isothiocyanate-labeled plasmid and Texas-Red-labeled chitosan. Several factors affect transfection activity and cell uptake, for example the molecular mass of chitosan, stoichiometry of complex, seriun concentration and the pH of the transfection medium. The level of transfection with plasmid-chitosan complexes was found to be highest when the molecular mass of chitosan was 40 or 84 kDa, the ratio of chitosan nitrogen to DNA phosphate was 5, and serum at pH 7.0 was 10%. Plasmid-chitosan complexes most likely condense to form large aggregates (5-8 p,m), which absorb to the cell surface. After this, plasmid-chitosan complexes are endocytosed, and accumulate in the nucleus [97]. 

Griffin DE III, Hill WE. 1978. In vitro breakage of plasmid DNA by mutagens and pesticides. Mutat Res 52 161-169. 

S., Hamer, )., Escobedo, J., Cohen, F., Radhakrishnan, R., Dwarki, V, and ZuCKERMANN, R.N. Lipitoids — novel cationic lipids for cellular delivery of plasmid DNA in vitro. Chem. Biol. 1998, 5, 345-354. 

FIG. 2 Atomic force microscopy of plasmid DNA at different stages of condensation with polycations. (a) Circular plasmid DNA (b) DNA condensed with poly-L-lysine (PLL, mol. wt. 4 kDa) at a DNA phosphate/lysine ratio of 2 1 (c) DNA condensed with PLL (mol. wt. 10 kDa) at a phos-phate/lysine ratio of 1 2 (d) toroid of DNA condensed with PLL-asialo-orosomucoid conjugate at a phosphate/lysine ratio of 1 6. [(a) and (d) from Ref. 80, copyright 1998 Oxford University Press (b) and (c) reprinted with permission from Ref. 66, copyright 1999 American Chemical Society.]... 

The size of plasmids used for transfection can vary considerably, but most plasmids are 4,000 to 10,000 base pairs in size. Despite their differences all plasmids face the same barriers when transfected. Transfected DNA has to cross the cell membrane or the endosomal membrane, it has to be transported into the nucleus, and it has to be protected against cellular nucleases and degradation... 

Fig. 3. Comparison of transfection efficiencies obtained using PolyFect Reagent, a dendrimer-based transfection reagent, and a calcium phosphate-mediated procedure. COS-7 and HeLa cells were transfected in srx-weU plates with a /3-galactosidase expression plasmid using the appropriate protocol. For the calcium phosphate-mediated transfection, 6 pg of plasmid DNA was used and the medium was changed after 5 h incubation. Transfections were performed in triplicate, and transfection efficiency was measured by monitoring the /3-galactosidase activity of extracts obtained from the transfected cells. The amoimt of /3-galactosidase activity in the extracts correlates with the transfection efficiency. Cells were harvested 48 h post-trans-fection...

Other pseudomonic acids (B, C, D) are also produced. Mupirocin is active predominantly against staphylococci and most streptococci, but Enterococcus faecalis and Gramnegative bacilli are resistant There is also evidence of plasmid-mediated mupirocin resistance in some chnical isolates of Staph, aureus. 

A third mechanism of plasmid transfer is by transformation, which is the ability of certain microorganisms to acquire naked DNA from the environment. This is limited to certain bacteria, notably Neisseria gonorrhoeae, which is naturally competent to acquire DNA in this manner. Neisseria gonorrhoeae strains have the ability to recognize DNA from their own species, and are thus selective in their acquisition of naked DNA from the environment. 

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