Secreted .alpha.-amylase as a reporter gene

Details Secreted .alpha.-amylase as a reporter gene Secreted .alpha.-amylase as a reporter gene

C - Chemistry, Metallurgy

12

N

C12N 15/85 (2006.01) C12N 9/26 (2006.01) C12N 15/56 (2006.01) C12N 15/62 (2006.01) C12Q 1/40 (2006.01) C12Q 1/68 (2006.01)

Patent

CA 2203613

The regulation of gene expression is of fundamental importance to all biological functions including adaptation to environmental conditions, cell division and differentiation, and the development of disease states such as cancer. The application of "reporter" gene systems, which allow changes in gene expression to be assayed quickly and easily, has contributed greatly to our understanding of the mechanisms involved in gene regulation. Reporter genes come in two basic varieties; those that encode intracellular enzymes, and those that code for secreted reporter proteins. Most reporter genes currently in use are of the intracellular variety that require manipulation of the cells, normally harvesting and lysis, prior to measurement of the reporter activity. In contrast, cells transformed with constructs that encode secreted reporter enzymes do not require manipulation or lysis of the cells for activity measurements. Instead, aliquots of the extracellular medium are harvested and assayed directly for enzyme activity. Thus, secreted reporter genes can be used to monitor gene expression in vitro, in vivo and ex vivo. The present invention relates to a reporter gene encoding the chicken a-amylase enzyme for use in transforming mammalian cells. The use of a-amylase as a secreted reporter gene is particularly attractive because the enzyme is extremely stable over a wide range of conditions, measurement of .alpha.- amylase activity is simple, quantitative, sensitive, safe and inexpensive, and the range of electrophoretically-distinguishable variants available allows the assays to be performed in virtually any host without interference from endogenous amylase activities.

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