4-5.2.1(b): Schematic process for screening libraries by Plaque hybridization.

Probes used for hybridization

Cloned DNA fragments can be used as probes in hybridization reactions if a cDNA clone is available. DNA or synthetic oligonucleotide probes can be used for identification of a clone from a genomic library instead of RNA probes, for example, to study the regulatory sequences which are not part of the cDNA clone. A common method of labeling probes is the incorporation of a radioactive or other marker into the molecule. A number of alternative labeling methods are also available that involve an amplification process to detect the presence of small quantities of bound probe and avoid the use of radioactivity. These methods involve the incorporation of chemical labels such as digoxigenin or biotin into the probe which can be detected with a specific antibody or the ligand streptavidin, respectively.

4-5.2.2. Screening by PCR

PCR screening is employed for the identification of rare DNA sequences in complex mixtures of molecular clones by increasing the abundance of a particular sequence. It is possible to identify any clone by PCR only if there is available information about its sequence to design suitable primers.