DNA from two or more sources is incorporated into a single recombinant molecule. To be usefulthe recombinant molecule must be replicated many times to provide material for analysis, sequencingetc. Producing many identical copies of the recombinant dna biology definition recombinant molecule is called cloning.
Cloning can be done in vitroby a process called the polymerase chain reaction PCR. Here, however, we shall examine how cloning is done in vivo. In every case, the recombinant DNA must be taken up by the cell in a form in which it can be replicated and expressed. This is achieved by incorporating the DNA in a vector. A number of viruses both bacterial and of mammalian cells can serve as vectors.
But here let us examine an example of cloning using E. Plasmids are molecules of DNA that are found recombinant dna biology definition bacteria separate from the bacterial chromosome. Plasmids are replicated by the same machinery that replicates the bacterial chromosome.
Some plasmids are copied at about the same rate as the chromosome, so a single cell is apt to have only a single copy of the plasmid. Other plasmids are copied at a high rate and a single cell may have 50 or more of them. Genes on plasmids with high numbers recombinant dna biology definition copies are usually expressed at high levels. In nature, these genes often encode proteins e. Plasmids enter the bacterial cell with relative ease. This occurs in nature and may account for the rapid spread of antibiotic resistance in hospitals and elsewhere.
Plasmids can be deliberately introduced into bacteria in the laboratory transforming the cell with the incoming genes. If you remove the two restriction enzymes and provide the recombinant dna biology definition for DNA ligase to do its work, the pieces of these plasmids can rejoin thanks to the complementarity of their sticky ends.
Some of these will not produce functional plasmids molecules with two or with no replication origin cannot function. One interesting possibility is the joining of the bp pAMP fragment with amp r and a replication origin with the bp pKAN fragment with kan r Sealed with DNA ligasethese molecules are functioning plasmids that are capable recombinant dna biology definition conferring resistance to both ampicillin and kanamycin. They are molecules of recombinant DNA. Because the recombinant dna biology definition origin, which enables the molecule to function as a plasmidwas contributed by pAMP, pAMP is called the vector.
If we treat any other sample of DNA, e. Mixed with EcoRI-treated plasmid and DNA ligase, a small number of the human molecules will become incorporated into the plasmid which can then be used to transform E. But how to detect those clones of E. The key is that the EcoRI site is within the kan r recombinant dna biology definition, so when a piece of human DNA is inserted there, the gene's function is destroyed.
All those clones that continue to grow on ampicillin but fail to grow on kanamycin here, clones 25and 8 have been transformed with a piece of human DNA. Electron micrograph of an E. The recombinant dna biology definition is a portion of a single DNA molecule containing over 4. The small circlets are plasmids.
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