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| Gene Cloning |
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| Gene cloning refers to the production of a large population of a DNA fragment in pure form. Gene cloning involves the formation of a recombinant DNA and its introduction into an appropriate host, such as Escherichia coli or Bacillus subtilis. The host used should be without plasmids. The walls of host bacteria are made permeable by treatment with calcium chloride or a lysozyme. The recombinant DNA is added to the culture in which such host bacteria are growing. The recombinant DNA is taken up by the bacteria along with the nutrients. It replicates whenever the bacterial cell divides. |
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| The bacterium E. coli divides every 22 minutes under favourable conditions, theoretically giving rise to a billion cells in less than 11 hours. This will produce a large clone of recombinant DNA. |
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| In some cases, the bacterial cells also transcribe the recombinant DNA molecules into mRNA, which is, in turn, translated into polypeptide chains. Hence, cloning can be employed not only to produce large quantities of the DNA sequence of an individual gene but also large amounts of the genes protein product as well. |
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| The process of gene cloning can be used for producing eukaryotic proteins in the bacterial cells, and for establishing a gene library. Following are some of the fields where gene cloning has found its application. |
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| Medical Utility |
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| Bacteria may be used as living laboratories for synthesising vitamins, hormones and antibiotics by introducing into them the genes which code for these substances along with the plasmids. |
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| Agricultural Utility |
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| The nitrogen fixing genes of bacteria may be transferred to the major food crops to boost food production without using expensive fertilizers. |
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| Gene Library |
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| It is a technique used for creating multiple copies of a given gene. A chromosome of an organism is broken into small fragments comprising of one or a few genes with the help of restriction endo-nucleases. Each fragment is joined to a suitable vehicle DNA, forming recombinant DNAs of different nature. These are then introduced into different host cells (bacterial, yeast, animal or plant) grown in culture. Each foreign DNA fragment replicates as its host cell divides. This produces several groups or clones of host cells, each containing a different fragment of the same foreign DNA. The various clones representing all the genes of an organism are referred to as a gene library of that organism library, a clone having a specific gene can be identified and this gene can be multiplied by growing the relevant clone in a culture for study, as and when needed. |
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| From a gene library, it is possible to identify a clone containing the specific gene. This gene can be multiplied by growing the relevant clone in a culture. The base sequence in this gene can be found, as the methods for this purpose are known. From the base sequence, the sequence of amino acids in a polypeptide can be worked out on the basis of the triplet code. |
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