( northern blot analysis or nuclease pro...)Transfection Reagent Selector Kit Handbook

northern blot analysis or nuclease protection assays to quantitate the specific mRNAs transcribed from the gene of interest. Since these procedures are time-consuming and inconvenient for multiple samples (resulting from multiple constructs), an alternative approach is to link the presumed cis-acting sequences from the gene of interest to the coding sequence of an unrelated reporter gene (see examples below) (3, 4). The reporter gene provides an indirect way of measuring how such regulatory sequences influence gene expression. Reporter genes are also useful in serving as controls. Transfection efficiencies between transfection experiments can be standardized by comparing the expression of the reporter gene product. Further information on genetic reporter systems can be obtained from current molecular biology manuals (3, 4).

In choosing a suitable reporter system, several considerations should be taken into account. First, the reporter gene should be absent from the cells used in the study or easily distinguished from the native form of the gene. Second, the assay for the reporter gene product should be quick, easy, sensitive, and inexpensive. In particular, a broad linear range is important to enable detection of both small and large changes in the reporter gene expression. Finally, the presence of the reporter gene should not affect the physiology of the cells being used.

Commonly-Used Reporters

Chloramphenicol acetyltransferase

The prokaryotic enzyme chloramphenicol acetyltransferase (CAT) is commonly used as a reporter. This enzyme catalyzes the transfer of acetyl-groups from acetyl-coenzyme A to ch
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