chapter  17
4 Pages

Differential Screening: Subtractive Libraries, AFLP-cDNA

Differential screening is a technique for cloning cDNAs corresponding to several specific mRNAs of a given tissue at a particular stage of development or under the effect of stress. The principle consists primarily of constructing a cDNA library from mRNA obtained by a treatment in which the specific cDNAs are to be isolated (e.g., leaves). Then, single-stranded (ss) cDNAs of the target tissue and another tissue (e.g., roots) are synthesized by reverse transcription of RNAs extracted from the two types of tissue. These cDNAs are then labelled to be used as probes. The library is then partly screened with the two probes. Some cDNA clones from the library will hybridize with the two probes: these correspond to mRNAs accumulated in the two types of tissue. In contrast, some cDNAs will hybridize only with the probe of the cDNAs from the target tissue: these correspond to specific mRNAs of the target tissue. These clones are then purified and characterized.