Subtractive Hybridization and Construction of cDNA Libraries
Genes that are differentially expressed both in time and space are the basis for how single cells, through the process of embryonic development, give rise to animals with an extraordinary diversity of cell types. As a first step in understanding differential gene expression, many researchers seek to identify those genes whose transcripts are temporally or spatially restricted to Particular cells, tissues, or embryonic stages. Although there are a variety of methods suitable for identifying moderately to highly expressed genes, the isolation of the most interesting class of mRNAs, those that are not abundant, but that may be cell-or tissue-specific, remains the most difficult task.
Several basic types of methods have been employed to identify low-abundance, tissue-specific transcripts. The more classical differential hybridization techniques (e.g., 1) are mostly limited to the detection of moderately abundant transcripts representing >0.05% of the mRNA population (2)....
KeywordsSodium Dodecyl Sulfate Isoamyl Alcohol Subtract cDNA Library Guanidine Thiocyanate Column Buffer
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