Woodruff Professor of Law; Alonzo L.
Natural cloning[ edit ] Cloning is a natural form of reproduction that has allowed life forms to spread for hundreds of millions of years. It is the reproduction method used by plantsfungiand bacteriaand is also the way that clonal colonies reproduce themselves.
Molecular cloning Molecular cloning refers to the process of making multiple molecules. Cloning is commonly used to amplify DNA fragments containing whole genesbut it can also be used to amplify any DNA sequence such as promotersnon-coding sequences and randomly fragmented DNA.
It is used in a wide array of biological experiments and practical applications ranging from genetic fingerprinting to large scale protein production. Occasionally, the term cloning is misleadingly used to refer to the identification of the chromosomal location of a gene associated with a particular phenotype of interest, such as in positional cloning.
In practice, localization of the gene to a chromosome or genomic region does not necessarily enable one to isolate or amplify the relevant genomic sequence.
To amplify any DNA sequence in a living organism, that sequence must be linked to an origin of replicationwhich is a sequence of DNA capable of directing the propagation of itself and any linked sequence. However, a number of other features are needed, and a variety of specialised cloning vectors small piece of DNA into which a foreign DNA fragment can be inserted exist that allow protein productionaffinity taggingsingle stranded RNA or DNA production and a host of other molecular biology tools.
Subsequently, a ligation procedure is used where the amplified fragment is inserted into a vector piece of DNA. The vector which is frequently circular is linearised using restriction enzymesand incubated with the fragment of interest under appropriate conditions with an enzyme called DNA ligase.
Following ligation the vector with the insert of interest is transfected into cells. A number of alternative techniques are available, such as chemical sensitivation of cells, electroporationoptical injection and biolistics.
Finally, the transfected cells are cultured. As the aforementioned procedures are of particularly low efficiency, there is a need to identify the cells that have been successfully transfected with the vector construct containing the desired insertion sequence in the required orientation.
Modern cloning vectors include selectable antibiotic resistance markers, which allow only cells in which the vector has been transfected, to grow. Nevertheless, these selection steps do not absolutely guarantee that the DNA insert is present in the cells obtained.
Further investigation of the resulting colonies must be required to confirm that cloning was successful. Cloning unicellular organisms[ edit ] Cloning cell-line colonies using cloning rings Cloning a cell means to derive a population of cells from a single cell.
In the case of unicellular organisms such as bacteria and yeast, this process is remarkably simple and essentially only requires the inoculation of the appropriate medium.
However, in the case of cell cultures from multi-cellular organisms, cell cloning is an arduous task as these cells will not readily grow in standard media. A useful tissue culture technique used to clone distinct lineages of cell lines involves the use of cloning rings cylinders.
At an early growth stage when colonies consist of only a few cells, sterile polystyrene rings cloning ringswhich have been dipped in grease, are placed over an individual colony and a small amount of trypsin is added.
Cloned cells are collected from inside the ring and transferred to a new vessel for further growth. Cloning stem cells[ edit ] Main article: Somatic-cell nuclear transfer Somatic-cell nuclear transferknown as SCNT, can also be used to create embryos for research or therapeutic purposes.
The most likely purpose for this is to produce embryos for use in stem cell research.Type or paste a DOI name into the text box. Click Go. Your browser will take you to a Web page (URL) associated with that DOI name. Send questions or comments to doi.
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