Normalization of cDNA Libraries
In a cellular transcriptome, the number of mRNA copies per gene may differ by several orders. In cDNA libraries, performed from mRNA, these proportions are the same. Normalization methods allow us to equalize numbers of gene’s copies in the library. Normalized cDNA libraries are used to discover new genes transcribed at relatively low levels or for functional screenings. Here, we observed different cDNA libraries normalization methods, which were based on hybridization (renaturation) of cDNA or DNA, or RNA. Also we described duplex-specific nuclease (DSN) normalization protocol – simple and effective cDNA libraries normalization method.
KeywordsGenetic regulation differentially expressed mRNA mRNA study poly(A) + RNA full-length cDNA incomplete cDNA cDNA representation transcript concentration normalized cDNA libraries poorly transcribed clone coverage saturating hybridization enzymatic removal physical separation biotinylation biotinylation of initial RNA frequent cutter duplex-specific nuclease (DSN) hydroxyapatite column plasmid vector phagemid vector first strand cDNA second strand cDNA immobilization
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