Methods for Generation and Analysis of Fluorescent Protein-Tagged Maize Lines
The use of fluorescent proteins to localize gene products in living cells has revolutionized cell biology. Although maize has excellent genetics resources, the use of fluorescent proteins in maize cell biology has not been well developed. To date, protein localization in this species has mostly been performed using immunolocalization with specific antibodies, when available, or by overexpression of fluorescent protein fusions. Localization of tagged proteins using native regulatory elements has the advantage that it is less likely to generate artifactual results, and also reports tissue-specific expression patterns for the gene of interest. Fluorescent protein tags can also be used for other applications, such as protein–protein interaction studies and purification of protein complexes. This chapter describes methods to generate and characterize fluorescent protein-tagged maize lines driven by their native regulatory elements.
Keywords:Confocal microscopy Gateway cloning™ Green fluorescent protein (GFP) Yellow FP (YFP) Red FP (RFP) Maize Native regulatory elements Protein localization Tissue-specific expression Triple-template PCR (TTPCR)
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