Isolating the clone

In general, cloning is undertaken in order to obtain the clone of one particular gene or DNA sequence of interest. The next step after cloning, therefore, is to find and isolate that clone among other members of the library. If the library encompasses the whole genome of an organism, then somewhere within that library will be the desired clone. There are several ways of finding it, depending on the specific gene concerned. Most commonly, a cloned DNA segment that shows homology to the sought gene is used as a probe. For example, if a mouse gene has already been cloned, then that clone can be used to find the equivalent human clone from a human genomic library. Bacterial colonies constituting a library are grown in a collection of Petri dishes. Then a porous membrane is laid over the surface of each plate, and cells adhere to the membrane. The cells are ruptured, and DNA is separated into single strands—all on the membrane. The probe is also separated into single strands and labeled, often with radioactive phosphorus. A solution of the radioactive probe is then used to bathe the membrane. The single-stranded probe DNA will adhere only to the DNA of the clone that contains the equivalent gene. The membrane is dried and placed against a sheet of radiation-sensitive film, and somewhere on the films a black spot will appear, announcing the presence and location of the desired clone. The clone can then be retrieved from the original Petri dishes.

Gene Cloning Introduction, Creating the clone