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Molecular cloning takes advantage of the fact that the chemical structure of DNA is fundamentally the same in all living organisms. Therefore, if any segment of DNA from any organism is inserted into a DNA segment containing the molecular sequences required for DNA replication, and the resulting recombinant DNA is introduced into the organism from which the replication sequences were obtained ...
Colony hybridization is a method of selecting bacterial colonies with desired genes through a straightforward cloning and transfer process. [1] The genes of interest have been added to a bacterial plasmid previously through recombination , allowing genes from other organisms to be analyzed within a bacterial colony.
A bacterial artificial chromosome (BAC) is a DNA construct, based on a functional fertility plasmid (or F-plasmid), used for transforming and cloning in bacteria, usually E. coli. [ 1 ] [ 2 ] [ 3 ] F-plasmids play a crucial role because they contain partition genes that promote the even distribution of plasmids after bacterial cell division.
Use the enzyme DNA ligase to seal the DNA fragments into the vector. This creates a large pool of recombinant molecules. These recombinant molecules are taken up by a host bacterium by transformation, creating a DNA library. [9] [10] Below is a diagram of the above outlined steps. Genomic Library Construction
Transformation is one of three forms of horizontal gene transfer that occur in nature among bacteria, in which DNA encoding for a trait passes from one bacterium to another and is integrated into the recipient genome by homologous recombination; the other two are transduction, carried out by means of a bacteriophage, and conjugation, in which a ...
Bacterial transformation involves moving a gene from one bacteria to another. It is integrated into the recipients plasmid. and can then be expressed by the new host. Transformation is the direct alteration of a cell's genetic components by passing the genetic material through the cell membrane.
A cloning vector is a DNA molecule that carries foreign DNA into a host cell, where it replicates, producing many copies of itself along with the foreign DNA. There are many types of cloning vectors such as plasmids and phages. In order to carry out recombination between vector and the foreign DNA, it is necessary the vector and DNA to be ...
Usually the ultimate aim of expression cloning is to produce large quantities of specific proteins.To this end, a bacterial expression clone may include a ribosome binding site (Shine-Dalgarno sequence) to enhance translation of the gene of interest's mRNA, a transcription termination sequence, or, in eukaryotes, specific sequences to promote the post-translational modification of the protein ...