Cassette plasmids/genes

The first of these sugar cassette plasmids, pLN2 [67], contains seven genes (oleVWUYLSE) from the oleandomycin pathway of S. antibioticus involved in the biosynthesis of NDP-L-oleandrose 20. In order to facilitate the expression and exchange of genes on the cassette plasmid, another plasmid, pFL942, which... 

Figure 2. A) Genetic organization of the pKAl catabolic plasmid. Genes of the upper naphthalene regulatory system encode for proteins that mediate the conversion of naphthalene to salicylate. Salicylate is then further degraded to TCA cycle intermediates. B) In Pseudomonas fluorescens HK44, genes within the lower pathway were replaced with genes of the lux cassette to produce a bioluminescent bioreporter sensitive to naphthalene and salicylate.

FIGURE 5.3.5 Enhancement of lycopene accumulation in . coZi by over-expression of DXS. Lycopene accumulation (left) is enhanced (right) when E. coli cells carrying a carotenoid pathway gene cassette (+EIB) are further transformed with a dxs gene on a multicopy plasmid (+E1B +dxs). Lycopene hyperaccumulation was demonstrated by Matthews and Wurtzel. ... 

DNA construct will often contain an effect gene and a selectable marker gene (such as antibiotic or herbicide resistance), both of which are bracketed by promoter and terminator sequences. A plasmid vector carries this cassette of genetic information into the plant genome by one of the above methods. 

Tennstedt T, Szczepanowski R, Braun S et al (2003) Occurrence of integron-associated resistance gene cassettes located on antibiotic resistance plasmids isolated from a waste water treatment plant. EEMS Microbiol Ecol 45(3) 239-252... 

The next challenge is to get the desired gene into the new cell. The target cell may be a bacterium, a yeast cell, or a cell from an insect, plant, or mammal. Scientists use delivery systems, called vectors, suited for the cell type, to get the combination of gene and promoter (sometimes called a cassette ) into the target cell, so that DNA will be copied each time the cell divides. The most commonly used vector to get a gene into bacteria is a plasmid, a small circular piece of DNA that is copied every time the bacterium divides into two, though it does not become part of the bacterial DNA. Plasmids have been developed that work as vectors for yeast, plant, and mammal cells. 

Many destination vectors are commercially available. However, it is also possible to construct a destination vector that is suitable for individual experiments. To constrnct snch vectors, one should insert the rr ffi-containg cassette (commercially available from Invitrogen) into the appropriate locns of a Gateway-incompatible vector. The resultant plasmids can be amplified only in the DB3.I strain, due to the toxic ccd gene (7). 

Nonviral vector systems are usually either composed of a plasmid based expression cassette alone ( naked DNA), or are prepared with a synthetic amphipathic DNA-complexing agent (84, 88). Gene delivery systems based on nonviral vectors mainly comprise cationic liposomes, DNA-polymer-protein complexes, and mechanic administration of naked DNA. An idealized/optimized multifunctional nonviral gene delivery system is depicted in Figure 13.4. 

Keil, S. Keil, H. (1992). Construction of a cassette enabling regulated gene expression in the presence of aromatic hydrocarbons. Plasmid, 27, 191-9-... 

Fig. 1. Structure of the expression cassette of a typical bacterial expression plasmid designed to produce an antibody Fab linked to a foreign gene (FG), pelB leader sequences direct penplasmic secretion of the nascent chains. The remainder of the plasmid contains the ampiciilin resistance gene and a plasmid origin of replication.

Naked plasmid DNA was not only trapped in the cytoplasm around the site of microinjection, as visualized by fluorescence in situ hybridization (FISH) or using FITC-labeled DNA, but was also eliminated rapidly at physiological temperature (Lechardeur et al., 1999). The disposal of the DNA was completely prevented when the cells were kept at 4°C (Lechardeur et al., 1999). A similar conclusion was reached by monitoring the amount of microinjected expression cassette by the polymerase chain reaction (PCR) (Pollard et al., 2001), suggesting that the metabolic instability of naked DNA contributes to the low efficiency of gene transfer (Lechardeur et al., 1999 Mirzayans et al, 1992 Neves etal., 1999 Pollard et al., 2001). 

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