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In addition, changes may be needed to the coding sequences as well, to optimize translation, make the protein soluble, direct the recombinant protein to the proper cellular or extracellular location, and stabilize the protein from degradation.In most cases, organisms containing recombinant DNA have apparently normal phenotypes. The host cells copy the vector DNA along with their own DNA, creating multiple copies of the inserted DNA. The vector DNA is isolated (or separated) from the host cells’ DNA and purified.
Construction of recombinant DNA, in which a foreign DNA fragment is inserted into a plasmid vector.
In this example, the gene indicated by the white color is inactivated upon insertion of the foreign DNA fragment.
Using recombinant DNA technology and synthetic DNA, literally any DNA sequence may be created and introduced into any of a very wide range of living organisms.
Proteins that can result from the expression of recombinant DNA within living cells are termed recombinant proteins.
promoter, translational initiation signal, and transcriptional terminator).
Specific changes to the host organism may be made to improve expression of the ectopic gene.
Formation of recombinant DNA requires a cloning vector, a DNA molecule that replicates within a living cell.
Vectors are generally derived from plasmids or viruses, and represent relatively small segments of DNA that contain necessary genetic signals for replication, as well as additional elements for convenience in inserting foreign DNA, identifying cells that contain recombinant DNA, and, where appropriate, expressing the foreign DNA.
In standard cloning protocols, the cloning of any DNA fragment essentially involves seven steps: (1) Choice of host organism and cloning vector, (2) Preparation of vector DNA, (3) Preparation of DNA to be cloned, (4) Creation of recombinant DNA, (5) Introduction of recombinant DNA into the host organism, (6) Selection of organisms containing recombinant DNA, and (7) Screening for clones with desired DNA inserts and biological properties.
Following transplantation into the host organism, the foreign DNA contained within the recombinant DNA construct may or may not be expressed.