Plasmids
are defined as autonomous elements, whose genomes exist in the cell as
extrachromosomal units. They are self replicating circular (only rarely linear)
duplex DNA molecules, which are maintained in a characteristics number of
copies in a bacterial cell, yeast cell or even in organelles found in
eukaryotic cells. These plasmids can be single
copy plasmids that are maintained as one plasmid DNA per cell or multicopy plasmids, which are
maintained as 10-20 genomes per cell. There are also plasmids, which are under relaxed replication control, thus
permitting their accumulation in very large numbers (up to 1000 copies per
cell). These are the plasmids which are used as cloning vectors, due to their
increased yield potential.
Circular
plasmid DNA which is used as a vector, can be cleaved at one site with the help
of a restriction enzyme to give a linear DNA molecule. A foreign DNA segment
can now be inserted, by joining the ends of broken circular DNA to the two ends
of foreign DNA, thus regenerating a bigger circular DNA molecule that can now
be separated by gel electrophoresis on the basis of its size. Selection of
chimeric DNA is also facilitated by the resistance genes, which the plasmid may
carry against one or more antibiotics. If a plasmid has two such genes
conferring resistance against two antibodies and if the foreign DNA insertion
site lies within one of these two genes, then the chimeric vector loses
resistance against one antibiotic. In such a situation, the parent vector in
bacterial cells can be selected by resistance against two antibodies and the
chimeric DNA can be selected by retention of resistance against only one of the
two antibiotics.
Authored and Published by;
Raj Abhisek Panda
