Bimolecular fluorescence complementation (BiFC) assay for protein-protein interaction in onion cells using the helios gene gun.
Investigation of gene function in diverse organisms relies on knowledge of how the gene products interact with each other in their normal cellular environment. The Bimolecular Fluorescence Complementation (BiFC) Assay(1) allows researchers to visualize protein-protein interactions in living cells and has become an essential research tool. This assay is based on the facilitated association of two fragments of a fluorescent protein (GFP) that are each fused to a potential interacting protein partner. The interaction of the two protein partners would facilitate the association of the N-terminal and C-terminal fragment of GFP, leading to fluorescence. For plant researchers, onion epidermal cells are an ideal experimental system for conducting the BiFC assay because of the ease in obtaining and preparing onion tissues and the direct visualization of fluorescence with minimal background fluorescence. The Helios Gene Gun (BioRad) is commonly used for bombarding plasmid DNA into onion cells. We demonstrate the use of Helios Gene Gun to introduce plasmid constructs for two interacting Arabidopsis thaliana transcription factors, SEUSS (SEU) and LEUNIG HOMOLOG (LUH)(2) and the visualization of their interactions mediated by BiFC in onion epidermal cells.
Hollender
Courtney A
CA
Dept. Of Cell Biology and Molecular Genetics, University of Maryland, MD, USA.
Liu
Zhongchi
Z
D017360:Arabidopsis / Q000235:genetics; D029681:Arabidopsis Proteins / Q000235:genetics; D019470:Biolistics / Q000295:instrumentation / Q000379:methods*; D008163:Luminescent Measurements / Q000379:methods*; D008856:Microscopy, Fluorescence / Q000295:instrumentation / Q000379:methods; D019697:Onions / Q000235:genetics* / Q000378:metabolism; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics*; D025941:Protein Interaction Mapping / Q000379:methods*; D014157:Transcription Factors / Q000235:genetics
D029681:Arabidopsis Proteins; C417768:LUG protein, Arabidopsis; C445880:SEUSS protein, Arabidopsis; D014157:Transcription Factors