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mRNA differential display(图)

标签: mRNA differential display

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All living organisms have thousands to tens of thousands of unique genes encoded in their genome, of which only a small fraction, perhaps 15%, are expressed in any individual cell. Therefore, it is the temporal and spatial regulation in gene expression that determines life processes. The course of normal cellular development as well as pathological changes that arise in diseases such as cancer are all believed to be driven by changes in gene expression. A pressing problem is to identify and characterize those genes that are differentially expressed in order to understand the molecular nature of disease state and subsequently, to devise rational therapies. Differential Display was invented in 1992 by Drs. Arthur Pardee and Peng Liang to allow rapid, accurate and sensitive detection of altered gene expression (Science. 1992, 257:967; U.S. Patent 5,262,311).

The mRNA Differential Display technology works by systematic amplification of the 3' terminal portions of mRNAs and resolution of those fragments on a DNA sequencing gel. Using anchored primers designed to bind to the 5' boundary of the poly-A tails for the reverse transcription, followed by PCR amplification with additional upstream primers of arbitrary sequences, mRNA sub-populations are visualized by denaturing polyacrylamide electrophoresis (See schematic illustration of differential display). This allows direct side-by-side comparison of most of the mRNAs between or among related cells. The differential display method is thus far unique in its potential to visualize all the expressed genes in a eukaryotic cell in a systematic and sequence-dependent manner by using multiple primer combinations. More importantly, the new method enables the recovery of sequence information and the development of probes to isolate their cDNA and genomic DNA for further molecular and functional characterizations. Because of its simplicity, sensitivity, and reproducibility, the mRNA Differential Display method is finding wide-ranging and rapid applications in developmental biology, cancer research, neuroscience, pathology, endocrinology, plant physiology, and many other fields.

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