what is cloning?
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Answer:
clowning is the process of producing genetically identical individuals of an organisms either naturally or artificially..In Nature, many organisms produce clones through asexual reproduction. cloning in biotechnology refers to the process of creating clones of organisms or copies or cells or DNA fragments...
there are 3 types of artificial cloning-
- gene cloning
- reproductive cloning
- therapeutic cloning
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Cloning is the process of producing genetically identical individuals of an organism either naturally or artificially. In nature, many organisms produce clones through asexual reproduction. Cloning in biotechnology refers to the process of creating clones of organisms or copies of cells or DNA fragments (molecular cloning). Beyond biology, the term refers to the production of multiple copies of digital media or software
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 plants, fungi, and bacteria, and is also the way that clonal colonies reproduce themselves. Examples of these organisms include blueberry plants, hazel trees, the Pando trees, the Kentucky coffeetree, Myrica, and the American sweetgum.
Molecular cloning refers to the process of making multiple molecules. Cloning is commonly used to amplify DNA fragments containing whole genes, but it can also be used to amplify any DNA sequence such as promoters, non-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 replication, which 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 production, affinity tagging, single stranded RNA or DNA production and a host of other molecular biology tools.
Cloning of any DNA fragment essentially involves four steps
fragmentation - breaking apart a strand of DNA
ligation - gluing together pieces of DNA in a desired sequence
transfection – inserting the newly formed pieces of DNA into cells
screening/selection – selecting out the cells that were successfully transfected with the new DNA
Although these steps are invariable among cloning procedures a number of alternative routes can be selected; these are summarized as a cloning strategy.
Initially, the DNA of interest needs to be isolated to provide a DNA segment of suitable size. 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 enzymes, and 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, electroporation, optical 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. Additionally, the cloning vectors may contain colour selection markers, which provide blue/white screening (alpha-factor complementation) on X-gal medium. 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. This may be accomplished by means of PCR, restriction fragment analysis and/or DNA sequencing.
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