Fluorometric GUS Analysis for Transformed Plant Material
The use of reporter genes in transgenic plants provides an excellent opportunity to investigate the ways in which promoters and other regulatory elements regulate gene expression. Neomycin phosphotransferase II (1), chloramphenicol acetyltransferase (2), luciferase (3), and β-glucuronidase (GUS; 4) genes may each be used to provide some indications of the extent and sites of gene expression. Each reporter gene system has particular requirements for assaying gene expression and distinctive features. The neomycin phosphotransferase and chloramphenicol acetyltransferase systems require the use of radioisotopes or HPLC, whereas the luciferase system requires a luminometer or darkroom facilities. The GUS reporter gene system, in contrast, is quick, easy to use, sensitive, does not require radioisotopes, and is relatively inexpensive. Plant biotechnologists use the Escherichia coli GUSA gene in their assessments of reporter gene activity. The E. coli GUS has a monomeric mol wt of 68 kDa, and exists as a tetramer in vivo (5).
KeywordsNeomycin Phosphotransferase Reporter Gene Activity Methyl Umbelliferone Chloramphenicol Acetyltransferase Stop Buffer
- 5.Wilson, K. J., Giller, K. E., and Jefferson, R. A. (1991) Beta-Glucuronidase (GUS) operon fusions as a tool for studying plant-microbe interactions, in Advances in Molecular Genetics of Plant-Microbe Interactions (Hennecke, H. and Verma, D. P. S., eds.), Kluwer, The Netherlands, pp. 226–229.Google Scholar
- 6.Gallagher, S. R. (ed.) (1992) GUS Protocols, Academic, London.Google Scholar