Minicircles are small (~4 kb) circular replicons. They occur naturally in some

eukaryotic Eukaryotes () are organisms whose cells have a nucleus. All animals, plants, fungi, and many unicellular organisms, are Eukaryotes. They belong to the group of organisms Eukaryota or Eukarya, which is one of the three domains of life. Bacte ...

organelle

genomes In the fields of molecular biology and genetics, a genome is all the genetic information of an organism. It consists of nucleotide sequences of DNA (or RNA in RNA viruses). The nuclear genome includes protein-coding genes and non-coding g ...

. In the

mitochondria A mitochondrion (; ) is an organelle found in the Cell (biology), cells of most Eukaryotes, such as animals, plants and Fungus, fungi. Mitochondria have a double lipid bilayer, membrane structure and use aerobic respiration to generate adenosi ...

-derived kinetoplast of

trypanosomes Trypanosomatida is a group of kinetoplastid excavates distinguished by having only a single flagellum. The name is derived from the Greek ''trypano'' (borer) and ''soma'' (body) because of the corkscrew-like motion of some trypanosomatid species. ...

, minicircles encode

guide RNA A guide RNA (gRNA) is a piece of RNA that functions as a guide for RNA- or DNA-targeting enzymes, with which it forms complexes. Very often these enzymes will delete, insert or otherwise alter the targeted RNA or DNA. They occur naturally, se ...

s for

RNA editing RNA editing (also RNA modification) is a molecular process through which some cells can make discrete changes to specific nucleotide sequences within an RNA molecule after it has been generated by RNA polymerase. It occurs in all living organism ...

. In ''

Amphidinium ''Amphidinium'' is a genus of dinoflagellates. The type for the genus is '' Amphidinium operculatum'' Claparède & Lachmann. The genus includes the species '' Amphidinium carterae'' which is used as a model organism. As dinoflagellates, ''Amphi ...

'', the chloroplast genome is made of minicircles that encode chloroplast proteins.

''In vitro'' experimentally-derived minicircles

Minicircles are small (~4kb) circular

plasmid A plasmid is a small, extrachromosomal DNA molecule within a cell that is physically separated from chromosomal DNA and can replicate independently. They are most commonly found as small circular, double-stranded DNA molecules in bacteria; how ...

derivatives that have been freed from all

prokaryotic A prokaryote () is a single-celled organism that lacks a nucleus and other membrane-bound organelles. The word ''prokaryote'' comes from the Greek πρό (, 'before') and κάρυον (, 'nut' or 'kernel').Campbell, N. "Biology:Concepts & Connec ...

vector Vector most often refers to: *Euclidean vector, a quantity with a magnitude and a direction *Vector (epidemiology), an agent that carries and transmits an infectious pathogen into another living organism Vector may also refer to: Mathematic ...

parts. They have been applied as

transgene A transgene is a gene that has been transferred naturally, or by any of a number of genetic engineering techniques, from one organism to another. The introduction of a transgene, in a process known as transgenesis, has the potential to change th ...

carriers for the

genetic modification Genetic engineering, also called genetic modification or genetic manipulation, is the modification and manipulation of an organism's genes using technology. It is a set of technologies used to change the genetic makeup of cells, including t ...

of mammalian cells, with the advantage that, since they contain no bacterial DNA sequences, they are less likely to be perceived as foreign and destroyed. (Typical transgene delivery methods involve plasmids, which contain foreign DNA.) The smaller size of minicircles also extends their

cloning Cloning is the process of producing individual organisms with identical or virtually identical DNA, either by natural or artificial means. In nature, some organisms produce clones through asexual reproduction. In the field of biotechnology, cl ...

capacity and facilitates their delivery into cells. Their preparation usually follows a two-step procedure: * production of a 'parental plasmid' (bacterial plasmid with

inserts) in '' E. coli'' * induction of a site-specific recombinase at the end of this process but still in bacteria. These steps are followed by the ** excision of prokaryotic vector parts via two recombinase-target sequences at both ends of the insert ** recovery of the resulting minicircle (vehicle for the highly efficient modification of the recipient cell) and the miniplasmid by capillary gel electrophoresis (CGE) The purified minicircle can be transferred into the recipient cell by

transfection Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. It may also refer to other methods and cell types, although other terms are often preferred: " transformation" is typically used to des ...

lipofection Transfection is the process of deliberately introducing naked or purified nucleic acids into eukaryotic cells. It may also refer to other methods and cell types, although other terms are often preferred: " transformation" is typically used to desc ...

and into a differentiated tissue by, for instance, jet injection. Conventional minicircles lack an

origin of replication The origin of replication (also called the replication origin) is a particular sequence in a genome at which replication is initiated. Propagation of the genetic material between generations requires timely and accurate duplication of DNA by se ...

, so they do not replicate within the target cells and the encoded genes will disappear as the cell divides (which can be either an advantage or disadvantage depending on whether the application demands persistent or transient expression). A novel addition to the field are nonviral self-replicating minicircles, which owe this property to the presence of a S/MAR-Element. Self-replicating minicircles hold great promise for the systematic modification of stem cells and will significantly extend the potential of their plasmidal precursor forms ("parental plasmids"), the more as the principal feasibility of such an approach has amply been demonstrated for their plasmidal precursor forms.

References

{{Reflist Molecular genetics Applied genetics

''In vitro'' experimentally-derived minicircles

See also

References